Activity In Mixed Cultures Research Articles

PHA accumulation of a mixed microbial culture co-exists with ammonia partial nitritation

The nitrifying activity in mixed cultures enriched to accumulate polyhydroxyalkanoates (PHA) is in general inhibited by the addition of chemicals, such as allylthiourea (ATU). In the present study, a sequencing batch reactor was inoculated with a mixed microbial culture enriched in PHA-accumulating microorganisms, operating under complete aerobic conditions, feast/famine regime and with ATU addition. Genera Comamonas, Azoarcus and Thauera, all of them known as PHA-accumulating bacteria, were the dominant microbial populations present in the mixed culture. They showed a PHA storage capacity up to 60.0 wt%, as hydroxybutyrate (HB) and hydroxyvalerate (HV), in a HB:HV ratio of 63:37. After 20 days of operation, ATU addition was stopped and the reactor was run under the same operational conditions for 400 days. The development of partial nitritation activity (ammonia oxidation to nitrite) was observed, presumably due to free ammonia inhibition or the selected sludge retention time. Ammonium oxidizing bacteria (AOB) from genera Nitrosomonas, Nitrosococcus and Nitrosospira were detected, whereas the composition of the dominant PHA-accumulating microbial populations was the same as in the inoculum. This enriched mixed culture reached a maximum PHA accumulation of 59.2 wt%, with a HB:HV ratio of 59:41. Results showed the simultaneous occurrence of partial nitritation and the enrichment in PHA-accumulating microorganisms, which did not affect the maximum PHA storage capacity of the system.

Novel two-step process to improve efficiency of bio-oxidation of Axi high-sulfur refractory gold concentrates

In order to improve the bio-oxidation efficiency of Axi refractory gold concentrate, a two-step process including a high temperature chemical oxidation and a subsequent bio-oxidation, combined with pH control during the bio-oxidation step was used. The results revealed that the optimum mode was to maintain solution pH at 1.0-1.2 during the biological oxidation stage. Under this condition, the activity of mixed culture could be sustained and the formation of jarosite could be diminished, thus the oxidation efficiency was improved. The oxidation levels of iron and sulfur were improved by 12.50% and 15.49%, and the gold recovery was increased by 21.02%. Therefore, the two-step process combined with pH control is an effective method for oxidizing the biohydrometallurgical process of Axi gold concentrate, and it will have a broad prospect of application in dealing with complex refractory gold concentrate.

Sialic acid transporter NanT participates in Tannerella forsythia biofilm formation and survival on epithelial cells

Tannerella forsythia is a periodontal pathogen implicated in periodontitis. This gram-negative pathogen depends on exogenous peptidoglycan amino sugar N-acetylmuramic acid (NAM) for growth. In the biofilm state the bacterium can utilize sialic acid (Neu5Ac) instead of NAM to sustain its growth. Thus, the sialic acid utilization system of the bacterium plays a critical role in the growth and survival of the organism in the absence of NAM. We sought the function of a T. forsythia gene annotated as nanT coding for an inner-membrane sugar transporter located on a sialic acid utilization genetic cluster. To determine the function of this putative sialic acid transporter, an isogenic nanT-deletion mutant generated by allelic replacement strategy was evaluated for biofilm formation on NAM or Neu5Ac, and survival on KB epithelial cells. Moreover, since T. forsythia forms synergistic biofilms with Fusobacterium nucleatum, co-biofilm formation activity in mixed culture and sialic acid uptake in culture were also assessed. The data showed that the nanT-inactivated mutant of T. forsythia was attenuated in its ability to uptake sialic acid. The mutant formed weaker biofilms compared to the wild-type strain in the presence of sialic acid and as co-biofilms with F. nucleatum. Moreover, compared to the wild-type T. forsythia nanT-inactivated mutant showed reduced survival when incubated on KB epithelial cells. Taken together, the data presented here demonstrate that NanT-mediated sialic transportation is essential for sialic acid utilization during biofilm growth and survival of the organism on epithelial cells and implies sialic acid might be key for its survival both in subgingival biofilms and during infection of human epithelial cells in vivo.

Characterisation of the physical and chemical properties influencing bacterial epibiont communities on benthic gelatinous egg masses of the pulmonate Siphonaria diemenensis

The ability of sessile benthic egg masses to deter or prevent epibiosis is essential to the success of species that employ this life-history strategy. This study characterised the physical structure and bacterial communities on the surface of egg masses from the Siphonariid mollusc Siphonaria diemenensis (Quoy & Gaimard, 1833). Egg masses at the veliger stage of development were collected from two intertidal sites in the Gulf St. Vincent, South Australia. Physical structure was assessed using a combination of light microscopy and scanning electron microscopy. Egg mass surfaces were characterised by wave-like elevations 1–3μm apart, fouled only by cocci, and longitudinal ridges (5–20μm) fouled by a diversity of microorganisms and dense exopolymeric substance. Bacteria from the surface of egg masses and adjacent rock substratum were then isolated using standard culture procedures. The biochemical profiles of the isolates were used, along with Gram stain and visual morphological observations, to identify the bacteria. Eight species of bacteria were isolated and the composition of culturable epibiont communities from the egg mass was found to be significantly different from those found on the adjacent substrata. One species of bacterium on egg masses exhibited antibacterial activity in mixed culture and was identified as Bacillus psychrodurans using PCR of the partial 16S rRNA gene and sequence alignment on the GenBank database. Chemical extraction was performed on ‘clean’ and ‘fouled’ eggs and antibacterial activity was assessed against the marine pathogen Vibrio harveyi using the disc diffusion assay. Extracts from the cleaned egg masses were found to inhibit the growth of V. harveyi, whilst the fouled egg masses and extracts from the epibionts showed no antibacterial activity. However, extracts from the supernatant and cell pellet from the cultured B. psychrodurans exhibited antibacterial activity against V. harveyi and two human pathogens, Staphylococcus aureus and Escherichia coli. The results obtained in this study suggest that the surfaces of S. diemenensis egg masses are selective towards coccoid bacteria, which may result from a combination of physical structure and chemical antimicrobial properties, with further competitive interactions possibly occurring between the epibionts post settlement.

Multi-enzyme production by pure and mixed cultures of Saccharomyces and non-Saccharomyces yeasts during wine fermentation

Saccharomyces and non-Saccharomyces yeasts release enzymes that are able to transform neutral compounds of grape berries into active aromatic compounds, a process that enhances the sensory attributes of wines. So far, there exists only little information about enzymatic activity in mixed cultures of Saccharomyces and non-Saccharomyces during grape must fermentations. The aim of the present work was to determine the ability of yeasts to produce extracellular enzymes of enological relevance (β-glucosidases, pectinases, proteases, amylases or xylanases) in pure and mixed Saccharomyces/non-Saccharomyces cultures during fermentation. Pure and mixed cultures of Saccharomyces cerevisiae BSc562, Hanseniaspora vinae BHv438 and Torulaspora delbrueckii BTd259 were assayed: 1% S. cerevisiae/99% H. vinae, 10% S. cerevisiae/90% H. vinae, 1% S. cerevisiae/99% T. delbrueckii and 10% S. cerevisiae/90% T. delbrueckii. Microvinifications were carried out with fresh must without pressing from Vitis vinifera L. c.v. Pedro Jiménez, an autochthonous variety from Argentina. Non-Saccharomyces species survived during 15–18days (BTd259) or until the end of the fermentation (BHv438) and influenced enzymatic profiles of mixed cultures. The results suggest that high concentrations of sugars did not affect enzymatic activity. β-Glucosidase and pectinase activities seemed to be adversely affected by an increase in ethanol: activity diminished with increasing fermentation time. Throughout the fermentation, Saccharomyces and non-Saccharomyces isolates assayed produced a broad range of enzymes of enological interest that catalyze hydrolysis of polymers present in grape juice. Vinifications carried out by a pure or mixed culture of BTd259 (99% of T. delbrueckii) showed the highest production of all enzymes assayed except for β-glucosidase. In mixed cultures, S. cerevisiae outgrew H. vinae, and T. delbrueckii was only detected until halfway the fermentation process. Nevertheless, their secreted enzymes could be detected throughout the fermentation process.Our results may contribute to a better understanding of the microbial interactions and the influence of some enzymes on vinification environments.

Cognitive Decline in Alzheimer's Disease Is Associated with Selective Changes in Calcineurin/NFAT Signaling

Upon activation by calcineurin, the nuclear factor of activated T-cells (NFAT) translocates to the nucleus and guides the transcription of numerous molecules involved in inflammation and Ca(2+) dysregulation, both of which are prominent features of Alzheimer's disease (AD). However, NFAT signaling in AD remains relatively uninvestigated. Using isolated cytosolic and nuclear fractions prepared from rapid-autopsy postmortem human brain tissue, we show that NFATs 1 and 3 shifted to nuclear compartments in the hippocampus at different stages of neuropathology and cognitive decline, whereas NFAT2 remained unchanged. NFAT1 exhibited greater association with isolated nuclear fractions in subjects with mild cognitive impairment (MCI), whereas NFAT3 showed a strong nuclear bias in subjects with severe dementia and AD. Similar to NFAT1, calcineurin-Aalpha also exhibited a nuclear bias in the early stages of cognitive decline. But, unlike NFAT1 and similar to NFAT3, the nuclear bias for calcineurin became more pronounced as cognition worsened. Changes in calcineurin/NFAT3 were directly correlated to soluble amyloid-beta (Abeta((1-42))) levels in postmortem hippocampus, and oligomeric Abeta, in particular, robustly stimulated NFAT activation in primary rat astrocyte cultures. Oligomeric Abeta also caused a significant reduction in excitatory amino acid transporter 2 (EAAT2) protein levels in astrocyte cultures, which was blocked by NFAT inhibition. Moreover, inhibition of astrocytic NFAT activity in mixed cultures ameliorated Abeta-dependent elevations in glutamate and neuronal death. The results suggest that NFAT signaling is selectively altered in AD and may play an important role in driving Abeta-mediated neurodegeneration.

Unraveling microbial interactions in food fermentations: from classical to genomics approaches.

Fermentation, the microbial degradation of organic compounds without net oxidation, is an important process in the global carbon cycle and is also exploited worldwide for the production and preservation of food. It is one of the oldest food-processing technologies known, with some records dating back to 6,000 b.c. (50). The link between food and microbiology was laid by Pasteur, who found that yeasts were responsible for alcoholic fermentation (106). Since that discovery, scientific and industrial interests in food microbiology started to grow and continue to increase today. The number of food products that rely on fermentation in one or more steps of their production is tremendous. They form an important constituent of the daily diet and rank among the most innovative product categories in the food industry. Most of the important microorganisms applied in the production of fermented foods have been studied for decades, yielding a wealth of information on their physiology and genetics in relation to product functionalities, such as the development of flavor, taste, and texture. The recent emergence of genomics has opened new avenues for the systematic analysis of microbial metabolism and the responses of microorganisms to their environment. Additionally, genomics has boosted research on important food microbes (22, 90, 93). Much of this research focuses on the performance of a single strain, including its interactions with the food matrix. However, food fermentations are typically carried out by mixed cultures consisting of multiple strains or species. Population dynamics play a crucial role in the performance of mixed-culture fermentations, and for many years, studies on mixed-culture food fermentations have focused on analyzing population dynamics using classical and molecular methods. Many of these studies are mainly descriptive, and relatively little is known about the mechanisms governing population dynamics in general and the molecular interactions that occur between the consortium members in particular. The availability of genome sequences for several species that are of industrial importance as well as technological advances in functional genomics enable new approaches to study food microbiology beyond the single species level and allow an integral analysis of the interactions and metabolic activity in mixed cultures. Here we review the current knowledge on important food fermentation processes, focusing on the bacterial interactions. In addition, we illustrate how genomics approaches may contribute to the elucidation of the interaction networks between microbes, including interactions with the food environment. This information may find application in the industry through rational optimization and increased control over mixed-culture fermentations.

Probiotic activity of mixed cultures of kefir's lactobacilli and non-lactose fermenting yeasts.

Kefir is a fermented milk drink produced by lactic acid bacteria and lactose or non-lactose fermenting yeast, which are growing in a strong rel ationship, and is reported to have a unique taste a nd properties. During fermentation, peptides and exo-p olysaccharides with bioactive properties are formed . Moreover, by in vitro tests and animal trials, kefir and its constituent s have shown to have anticarcinogenic, antimutagenic, antiviral and antifungal properties .

Increase of acetate ester-hydrolysing esterase activity in mixed cultures of Saccharomyces cerevisiae and Pichia anomala

To examine the efficacy of mixed cultures with Saccharomyces cerevisiae and Pichia anomala on flavour profiles of alcoholic beverages, a Pichia mutant with low levels of ethyl acetate that negatively impact on the sensory quality was isolated. A petite mutant isolated from P. anomala NBRC 10213 treated with ethidium bromide had the lower activity of ethyl acetate-hydrolysing esterase (EAHase) than the wild-type in crude extracts. In the fermentation tests of pure cultures, the P. anomala mutant produced less ethanol, acetate and ethyl acetate than the wild-type. In mixed cultures with S. cerevisiae, the P. anomala mutant died quicker and produced lower amounts of ethyl acetate than the wild-type. Mixed cultures of S. cerevisiae and P. anomala showed higher activities of EAHase than pure culture of S. cerevisiae throughout the fermentation periods. The transition to the formation of acetate esters was considerably analogous to the transition to the activity of acetate ester-hydrolysing esterase with little time lag. The P. anomala mutant was superior to the wild-type in flavour profiles. The higher ethyl acetate concentrations formed mainly by P. anomala in mixed cultures are the primary stimulus for the EAHase in S. cerevisiae and the activity of acetate ester-hydrolysing esterase is crucial to the formation of acetate esters in mixed cultures of S. cerevisiae and P. anomala. An application of non-Saccharomyces yeast, P. anomala to enhance the sensory quality in alcoholic beverage and a mechanism of the formation of acetate esters in mixed cultures with S. cerevisiae and P. anomala were offered.

Cholesterol deficiency increases the vulnerability of hippocampal glia in primary culture to glutamate-induced excitotoxicity

Cholesterol, a molecule critical for cellular function, is found in particular high concentration in the brain and has been implicated to synaptic plasticity and neuronal regeneration. This study was undertaken to investigate the mechanism by which cholesterol shortage modulates glutamate (Glu)-induced excitotoxicity in hippocampal cell cultures. A combined treatment of lovastatin and β-cyclodextrin reduced cellular content of cholesterol while having no significant effect on cell viability in neuron/glia mixed cultures. The experimental manipulation, nonetheless, exacerbated Glu-induced membrane damage and loss of mitochondrial activity in mixed cultures. Analysis of [ 3 H ]thymidine incorporation revealed cholesterol deficiency impaired cell proliferation in mixed cultures after Glu exposure, indicating considerable loss of glia. Indeed, it was found that cholesterol deprivation potentiated the release of lactate dehydrogenase (LDH) and the impairment in mitochondrial reduction of WST-1 reagent in astrocyte-enriched cultures subjected to Glu exposure. The detrimental effect of cholesterol shortage, nevertheless, was not observed in cultured neurons. Notably, the pretreatment of lovastatin and β-cyclodextrin caused a decrease in the content of cellular LDH while having no effect on cell cycle profile and cellular activity of WST-1 reduction in astrocyte-enriched cultures. In contrast, removal of cholesterol had no effect on LDH content in neuron-enriched cultures. It is concluded that the differential vulnerability of cholesterol-depleted neural cells to excitotoxic damage may, in part, be ascribed to cholesterol shortage destabilizing the plasma membrane of astrocytes, thus rendering them less capable of withstanding Glu insult.

Transforming growth factor alpha-induced expression of type 1 plasminogen activator inhibitor in astrocytes rescues neurons from excitotoxicity.

Although transforming growth factor (TGF)-alpha, a member of the epidermal growth factor (EGF) family, has been shown to protect neurons against excitotoxic and ischemic brain injuries, its mechanism of action remains unknown. In the present study, we used in vitro models of apoptotic or necrotic paradigms demonstrating that TGF-alpha rescues neurons from N-methyl-D-aspartate (NMDA)-induced excitotoxic death, with the obligatory presence of astrocytes. Because neuronal tissue-type plasminogen activator (t-PA) release was shown to potentiate NMDA-induced excitotoxicity, we observed that TGF-alpha treatment reduced NMDA-induced increase of t-PA activity in mixed cultures of neurons and astrocytes. In addition, we showed that although TGF-alpha induces activation of the extracellular signal-regulated kinases (ERKs) in astrocytes, it failed to activate p42/p44 in neurons. Finally, we showed that TGF-alpha, by an ERK-dependent mechanism, stimulates the astrocytic expression of PAI-1, a t-PA inhibitor, which mediates the neuroprotective activity of TGF-alpha against NMDA-mediated excitotoxic neuronal death. Taken together, we indicate that TGF-alpha rescues neurons from NMDA-induced excitotoxicity in mixed cultures through inhibition of t-PA activity, involving PAI-1 overexpression by an ERK-dependent pathway in astrocytes.

Comparison of Mixed Cultures Based on Activity

Biodegradation experiments using mixed bacterial cultures are difficult to compare because repeatable rates may not be produced in different experiments when cultures are normalized on a cell mass or number basis. A technique was therefore developed to quantify [14C]substrate-specific biodegradation activity of mixed cultures. The technique can be used to obtain a preliminary estimate of the substrate-degrading activity of a culture for more substantial experiments. The technique is applied to 1-naphthol-humic acid bioavailability experiments using both mixed and pure cultures to help isolate the impact of humic acid from differences between the cultures.

Surfactants in relation to bioremediation and wastewater treatment

Surfactants have been shown to be beneficial in various technological applications. Recently, a lot of effort has been focused on modelling experimentally observed surfactant-enhanced dissolution, desorption and biodegradation phenomena of highly apolar organic compounds. In addition, the capacity of surfactants to enhance dispersal and growth of specific bacteria in soil has been demonstrated. Also, small amounts of surfactant have been shown to highly improve the anaerobic digestion process. Finally, the environmental fate of surfactants has been a subject of high interest. Although most surfactants appear to be biodegradable, their complete mineralization relies on the activity of mixed cultures. Considerable concern has recently been expressed about relatively stable intermediates which accumulate in the environment.

Interferon-gamma and tumor necrosis factor-alpha treatment of ex vivo human carcinoma cells potentiates their interaction with allogeneic lymphocytes.

Short-term exposure of ex vivo carcinoma and sarcoma cells to IFN-gamma and TNF-alpha induced or elevated to detectable levels the surface expression of MHC class I, class II, and ICAM-1 (CD54), but only rarely the B7 (CD80) molecules. The cytokine-treated tumor cells interacted more efficiently with allogeneic blood lymphocytes collected from healthy donors compared with untreated cells. This was demonstrated (1) by the induction of DNA synthesis and generation of cytotoxic activity in mixed cultures and (2) by the elevated susceptibility to the cytotoxic effectors. Although the cytokine-induced increase in MHC and ICAM-1 on the low-expressor tumors were probably important to the interaction with lymphocytes, it is likely that other properties were also induced that contributed to the phenomenon. This was indicated by the results obtained with several tumors that expressed indigenously high levels of these molecules but reacted with the allogeneic lymphocytes only or more efficiently after treatment with IFN-gamma and TNF-alpha. In these experiments B7 expression did not influence the efficiency of interactions between lymphocyte and tumor cells. The results also showed that, under the conditions used, the untreated tumor cells that did not activate allogeneic lymphocytes were sensitive to appropriately activated effectors. Thus the afferent and efferent arms of lymphocyte-tumor cell interactions appeared to have different requirements.

Nitrogenase activity in mixed cultures of Azospirillum with other bacteria

Summary Department of General Microbiology, Institute of Microbiology, Federal University of Rio de Janeiro, Brazil Fifty Gram-negative and 29 Gram-positive bacteria were tested in co-cultures in nitrogen-free, semisolid medium with either Azospirillum brasilense Sp7 or Azospirillum lipoferum Br17. Among the 79 bacteria examined, 12 Gram-negative and 7 Gram-positive did not exert any effect on the nitrgenase activity either in A. brasilense or A. lipoferum. The other bacteria stimulated or inhibited the nitrogenase either in one or both Azospirillum species. In several cases the same accompanying bacterium affected in a different way the two species of Azospirillum tested, stimulating one and inhibiting the other or acting on one, but not affecting the other. The active factors, produced by the accompanying bacteria, seem to be of oligodynamic nature. The capacity of producing these factors seems to be an individual pattern of particular strains, and no correlation was found between the taxonomical position of the accompanying bacterium and its manner of affecting Azospirillum.

Synthesis of Amylases by Mixed Cultures of Different Strains of Moulds

AbstractTwenty mould strains belonging to 13 species were used to determine the effect of mixed cultures on the synthesis of amylases. In test‐tube microcultures on a rotary shaker 110 pairs of combinations of these strains were studied. In 73 cases they dominated in their amylolytic activity over the weaker strain components in the culture, and in 21 cases also over stronger amylolytic components. Among six strain pairs selected for a more accurate evaluation all showed a higher glucoamylase activity in mixed cultures in comparison with an amylolytically less active component of the culture, and in four cases also in relation to a more active component.

Rapid Identification of Enterobacteriaceae

β‐glucuronidase activity is an exclusive characteristic of E. coli and some shigellae among Enterobacteriaceae and Vibrionaceae. An agar medium (PGUA agar) which permits the detection of bacteria with β‐glucuronidase activity in mixed cultures was evaluated as a primary culture medium for clinical samples of urine. The medium was selective for enterobacteria and yielded significantly higher recoveries than MacConkey agar. Based on the examination of 3,460 urine samples, it was found that the use of the PGUA agar has several advantages over conventional methods: 1) 94% of all E. coli cultures could be identified on the basis of their appearance on the primary plates; 2) The use of the PGUA method did not result in any misidentifications as compared to 1% of cultures misidentified by the conventional procedure; 3) Approximately one‐half of the urine samples which contained E. coli as the sole organism could be reported following the reading of primary culture plates; 4) The application of the PGUA medium resulted in a 46% reduction in the cost of media employed and a 67% reduction in the time required for the processing of urine samples.