Biofilm formation and chemostat dynamics: Pure and mixed culture considerations

Time-dependent biofilm formation effects on continuous fermenter operation are modelled here in general for a mixed culture of N different microorganisms growing on a single substrate. Dynamic computer solutions are detailed for two versions of the general model: a pure culture and a simple two-cell mixed culture. Pure culture model predictions compare favorably with two pure culture experiments in the literature where significant biofilm formation was noted. A mixed culture of one microbe (C(1)) having a higher growth rate than a second microbe (C(2)) is simulated for two hypothetical scenarios of microbe C(2) having different magnitudes of cell deposition rate. Biofilm effects on the estimation of kinetic and stoichiometric parameters in both model versions, plus the impact of biofilms on mixed culture dynamics, are discussed.     

Isolation and characterization of a symbiotic cellulolytic mixed bacterial culture

Summary By using batch-culture enrichment techniques a mixed culture of two bacterial spe cies identified as Cellulomonas flavigena and Xanthomonas sp was isolated. The capacity of both bacteria to grow as pure cultures in a min eral medium with alkaline pretreated sugar cane bagasse or cellobiose was tested. C. flavigena as pure culture was able to grow on both substrates only when yeast extract or biotin and thiamine were added to the culture medium, while Xanthomonas sp. could not grow on sugar cane ba gasse, but assimilated cellobiose if yeast extract was supplied. However, both bacteria in mixed culture grew very well on both substrates and did not require any growth factor. It was concluded that the interaction was favourable to both species. The mixed culture had the capacity to degrade a number of different agricul tural wastes and to use them as the sole carbon and energy source for the production mainly of biomass. More than 80% of pineapple bagasse, without chemical pretreatment, was used up by the microbial system.     

Effects of environmental settings on MTBE removal for a mixed culture and its monoculture isolation

A mixed culture was utilized to evaluate methyl tert-butyl ether (MTBE) removal under various conditions and to isolate a MTBE-degrading pure culture. The results showed that high MTBE removal efficiencies can be reached even in the presence of other substrates. The biodegradation sequence of the target compounds by the mixed culture, in order of removal rate, was toluene, ethyl benzene, p-xylene, benzene, MTBE, ethyl ether, tert-amyl methyl ether, and ethyl tert-butyl ether. In addition, preincubation of the mixed cultures with benzene and toluene showed no negative effect on MTBE removal; on the contrary, it could even increase the degradation rate of MTBE. The kinetic behavior showed that the maximum specific growth rate and the saturation constant of the mixed culture degrading MTBE are 0.000778 h⁻¹ and 0.029 mg l⁻¹, respectively. However, a high MTBE concentration (60 mg l⁻¹) was slightly inhibiting to the growth of the mixed culture. The pure culture isolated from the enrichments in the bubble-air bioreactor showed better efficiency in MTBE removal than the mixed culture; whereas, tert-butyl alcohol was formed as a metabolic intermediate during the breakdown of MTBE.     

Growth kinetics of top and bottom cultures of Saccharomyces spp. in a chemostat using sugarbeet molasses

Growth kinetics were evaluated for three yeast strains of the genus Saccharomyces. Two topfloating strains, SF 115 and SF 116 and one flocculant yeast SF 104 were analyzed in pure and mixed cultures in 1-liter continuous fermentation experiments in a chemostat. Growth was monitored for 72 h at 30°C in a medium containing sugarbeet molasses and 1.0 g/liter each of NH₄H₂PO₄ and urea. SF 115 and SF 116 were found to have lower μ_max values of 0.290 and 0.296 h^?1, respectively, than SF 104, which had a μ_max of 0.364 h^?1. The two top-floating yeasts (SF 115 and SF 116) demonstrated greater affinity for the substrate and utilized substrates at a greater rate. They have K₈ values of 4.03 × 10^?3 M and 3.798 × 10^?3 M, respectively, compared to 9.06 × 10^?3 M for SF 104. A mixed culture of SF 116 and SF and SF 104 was found to have a μ_max of 0.426 h^?1 with a K_s of 6.924 × 10^?3 M. SF 115 grown in mixed culture with SF 104 exhibited a μ_max of 0.473 h^?1 with a K_s of 7.975 × 10^?3 M. In both cases, the SF 104 was the dominant microbe in mixed culture systems.     

A Microsensor Study of the Interaction between Purple Sulfur and Green Sulfur Bacteria in Experimental Benthic Gradients

Abstract The interaction between the purple sulfur bacterium Thiocapsa roseopersicina and the green sulfur bacterium Prosthecochloris aestuarii was studied in a gradient chamber under a 16-hours light-8-hours dark regime. The effects of interaction were inferred by comparing the final outcome of a mixed culture experiment with those of the respective axenic cultures using the same inoculation densities and experimental conditions. Densities of bacteria were deduced from radiance microprofiles, and the chemical microenvironment was investigated with O₂, H₂S, and pH microelectrodes. P. aestuarii always formed a biofilm below the maximal oxygen penetration depth and its metabolism was strictly phototrophic. In contrast, T. roseopersicina formed a bilayer in both the mixed and the axenic culture. The top layer formed by the latter organism was exposed to oxygen, and chemotrophic sulfide oxidation took place during the dark periods, while the bottom layer grew phototrophically during the light periods only. In the mixed culture, the relative density of P. aestuarii was lower than in the axenic culture, which reflects the effects of the competition for sulfide. However, the relative density of T. roseopersicina was actually higher in the mixed culture than in the corresponding axenic culture, indicating a higher growth yield on sulfide in the mixed culture experiment. Several hypotheses are proposed to explain the effects of the interaction. Received: 15 June 1998; Accepted: 18 January 1999     

Editing of an effector gene promoter sequence impacts plant‐Phytophthora interaction

Pathogen avirulence (Avr) effectors interplay with corresponding plant resistance (R) proteins and activate robust plant immune responses. Although the expression pattern of Avr genes has been tied to their functions for a long time, it is still not clear how Avr gene expression patterns impact plant‐microbe interactions. Here, we selected PsAvr3b, which shows a typical effector gene expression pattern from a soybean root pathogen Phytophthora sojae. To modulate gene expression, we engineered PsAvr3b promoter sequences by in situ substitution with promoter sequences from Actin (constitutive expression), PsXEG1 (early expression), and PsNLP1 (later expression) using the CRISPR/Cas9. PsAvr3b driven by different promoters resulted in distinct expression levels across all the tested infection time points. Importantly, those mutants with low PsAvr3b expression successfully colonized soybean plants carrying the cognate R gene Rps3b. To dissect the difference in plant responses to the PsAvr3b expression level, we conducted RNA‐sequencing of different infection samples at 24 h postinfection and found soybean immune genes, including a few previously unknown genes that are associated with resistance. Our study highlights that fine‐tuning in Avr gene expression impacts the compatibility of plant disease and provides clues to improve crop resistance in disease control management.     

Emulsification and degradation of "Bunker C" fuel oil by microorganisms

An enrichment culture procedure has been used to isolate mixed culture systems which grow upon “Bunker C” fuel oil. When inoculated into a mineral salts aqueous medium containing Bunker C oil, the mixed cultures initiate oil emulsification. Emulsification usually is observed in 24–48 hr. The role of microbes in this emulsification will be discussed. It appears that certain metabolic products produced by the microbe possess properties of surfactants. Bacteria and fungi have been isolated which possess the ability to cause emulsification. Freeze-dried biomass is also capable of emulsifying oil. Chromatographic analyses of biodegraded Bunker C fuel oil show that microorganisms selectively metabolize the n-paraffin fraction.     

Maximum ingested food size in captive strepsirrhine primates: Scaling and the effects of diet

Little is known about ingested food size (V_b) in primates, even though this variable has potentially important effects on food intake and processing. This study provides the first data on V_b in strepsirrhine primates using a captive sample of 17 species. These data can be used for generating and testing models of feeding energetics. Strepsirrhines are of interest because they are hypometabolic and chewing rate and daily feeding time do not show a significant scaling relationship with body size. Using melon, carrot, and sweet potato we found that maximum V_b scales isometrically with body mass and mandible length. Low dietary quality in larger strepsirrhines might explain why V_b increases with body size at a greater rate than does resting metabolic rate. Relative to body size, V_b is large in frugivores but small in folivores; furthermore scaling slopes are higher in frugivores than in folivores. A gross estimate of dietary quality explains much of the variation in V_b that is not explained by body size. Gape adaptations might favor habitually large bites for frugivores and small ones for folivores. More data are required for several feeding variables and for wild populations. Am J Phys Anthropol 142:625–635, 2010. © 2010 Wiley‐Liss, Inc.     

The use of a mixed culture of Geotrichum candidum,Candida krusei and Hansenula anomala for microbial protein production from whiskey distillery spent wash

Summary The use of a mixed culture of Geotrichum candidum, Hansenula anomala and Candida krusei for microbial protein production and treatment of whiskey distillery spent wash was investigated in both batch and continuous culture. Although capable of assimilating the same substrate constituents in pure culture the organisms used different constituents for growth in mixed continuous culture, allowing a stable mixed population to be established. The relative proportions of the three organisms in the population was dependent on the dilution rate.The mixed culture did not give superior yields or productivities, but did give proportionally greater COD reduction and would be expected to bemore stable and resistant to contamination.At a dilution rate of 0.10 h^–1 the mixed culture gave a biomass yield of 4.8 g l^–1, containing 55% crude protein, with a COD reduction of 31.5%, while in mixed batch culture a biomass yield of 12.5 g l^–1, containing 48% crude protein, was obtained with a COD reduction of 54.9%.     

Use of Pseudomonas species producing phenazine-based metabolites in the anodes of microbial fuel cells to improve electricity generation

The rate of anodic electron transfer is one of the factors limiting the performance of microbial fuel cells (MFCs). It is known that phenazine-based metabolites produced by Pseudomonas species can function as electron shuttles for Pseudomonas themselves and also, in a syntrophic association, for Gram-positive bacteria. In this study, we have investigated whether phenazine-based metabolites and their producers could be used to improve the electricity generation of a MFC operated with a mixed culture. Both anodic supernatants obtained from MFCs operated with a Pseudomonas strain (P-PCA) producing phenazine-1-carboxylic acid (PCA) and those from MFCs operated with a strain (P-PCN) producing phenazine-1-carboxamide (PCN) exerted similarly positive effects on the electricity generation of a mixed culture. Replacing supernatants of MFCs operated with a mixed culture with supernatants of MFCs operated with P-PCN could double the currents generated. Purified PCA and purified PCN had similar effects. If the supernatant of an engineered strain overproducing PCN was used, the effect could be maintained over longer time courses, resulting in a 1.5-fold increase in the production of charge. Bioaugmentation of the mixed culture MFCs using slow release tubes containing P-PCN not only doubled the currents but also maintained the effect over longer periods. The results demonstrated the electron-shuttling effect of phenazine-based compounds produced by Pseudomonas species and their capacity to improve the performance of MFCs operated with mixed cultures. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The antibiotic action of methylarsenite is an emergent property of microbial communities

Arsenic is the most ubiquitous environmental toxin. Here, we demonstrate that bacteria have evolved the ability to use arsenic to gain a competitive advantage over other bacteria at least twice. Microbes generate toxic methylarsenite (MAs(III)) by methylation of arsenite (As(III)) or reduction of methylarsenate (MAs(V)). MAs(III) is oxidized aerobically to MAs(V), making methylation a detoxification process. MAs(V) is continually re‐reduced to MAs(III) by other community members, giving them a competitive advantage over sensitive bacteria. Because generation of a sustained pool of MAs(III) requires microbial communities, these complex interactions are an emergent property. We show that reduction of MAs(V) by Burkholderia sp. MR1 produces toxic MAs(III) that inhibits growth of Escherichia coli in mixed culture. There are three microbial mechanisms for resistance to MAs(III). ArsH oxidizes MAs(III) to MAs(V). ArsI degrades MAs(III) to As(III). ArsP confers resistance by efflux. Cells of E. coli expressing arsI, arsH or arsP grow in mixed culture with Burkholderia sp. MR1 in the presence of MAs(V). Thus MAs(III) has antibiotic properties: a toxic organic compound produced by one microbe to kill off competitors. Our results demonstrate that life has adapted to use environmental arsenic as a weapon in the continuing battle for dominance.     

Plant–microbe interactions promoting plant growth and health: perspectives for controlled use of microorganisms in agriculture

Plant-associated microorganisms fulfill important functions for plant growth and health. Direct plant growth promotion by microbes is based on improved nutrient acquisition and hormonal stimulation. Diverse mechanisms are involved in the suppression of plant pathogens, which is often indirectly connected with plant growth. Whereas members of the bacterial genera Azospirillum and Rhizobium are well-studied examples for plant growth promotion, Bacillus, Pseudomonas, Serratia, Stenotrophomonas, and Streptomyces and the fungal genera Ampelomyces, Coniothyrium, and Trichoderma are model organisms to demonstrate influence on plant health. Based on these beneficial plant–microbe interactions, it is possible to develop microbial inoculants for use in agricultural biotechnology. Dependent on their mode of action and effects, these products can be used as biofertilizers, plant strengtheners, phytostimulators, and biopesticides. There is a strong growing market for microbial inoculants worldwide with an annual growth rate of approximately 10%. The use of genomic technologies leads to products with more predictable and consistent effects. The future success of the biological control industry will benefit from interdisciplinary research, e.g., on mass production, formulation, interactions, and signaling with the environment, as well as on innovative business management, product marketing, and education. Altogether, the use of microorganisms and the exploitation of beneficial plant–microbe interactions offer promising and environmentally friendly strategies for conventional and organic agriculture worldwide.     

In vitro response as a reflection of genomic diversity in long-term cultures of Musa

Summary Ten commercial cultivars of Musa representing five different types of genomic constitutions were studied for in vitro multiplication through meristem culture. In addition, the effects of various genomic constitutions at different ploidy levels on growth and meristem proliferation in long-term cultures were analysed statistically. Plantlets were readily obtained by culturing the excised meristems on MS semisolid medium supplemented with IAA, IBA and BAP at various concentrations. The regenerative potential of all cultivars of Musa, irrespective of their genomes, remained unaffected in long-term culture, even after 28–30 months. The genomic influence on both the nature and rate of proliferative growth was evident. Statistical analysis revealed that the rates of meristem proliferation between different cultivars of the same passage and between different passages of the same cultivar were significantly different. Those cultivars having only an A genome showed a low rate of meristem proliferation, while under the same culture conditions, cultivars having one or two B genomes in addition to the A exhibited a very high rate.     

H-2 effects on cell-cell interactions in the response to single non-H-2 alloantigens

Individual mice were tested for their proliferative T-cell response to H-Y- and H-3-incompatible stimulator cells in secondary mixed lymphocyte culture. Responders expressing the H-2 ^bhaplotype were restricted in their response to stimulators presenting H-Y and H-3 in the context of H-2 ^b. Lymphocytes from individual B10 females proliferated in response to H-Y presented with I-A ^band D ^b. The ratio of I-A ^b/D ^b-restricted responses varied between individual responders, indicating significant qualitative variation between genetically identical responders. The majority of the proliferative response in all tested mice was restricted to the entire H-2 ^bhaplotype suggesting complementation of I-A ^b- and D ^b-region genes in presenting the H-Y antigen. Similar observations were made in the response of individual B10.LP mice to the H-3 antigen. H-3-specific, proliferating T cells were restricted to H-3 antigen presented with K ^bA^band D ^bwith significant variation between individuals in their preference for H-3 plus K ^bA^band D ^b. In contrast to the response to H-Y, the proliferative response to H-3 plus H-2 ^bcould be accounted for by the summation of the proliferative responses to H-3. plus K ^bA^band D ^b. These observations demonstrate that the proliferative response to non-H-2 H antigens in the context of I-region determinants is not a sine qua non for the T-cell response to these antigens. Further, the individual qualitative and quantitative variation observed with individual genetically identical mice has strong implications for our knowledge of intrastrain variation in immune responsiveness and the characterization of inbred strains for immune responsiveness.     

NITROGEN ASSIMILATION OF AN OCEANIC DIATOM IN NITROGEN-LIMITED CONTINUOUS CULTURE1

The oceanic diatom Thalassiosira pseudonana Hasle and Heimdal (formerly Cyclotella nana) was grown with 12L:12D illumination cycles in nitrogen-limited continuous culture with a mixture of ammonium and nitrate as the N source. Measurements included, at 3 different growth rates (degrees of N limitation), cell concentration, cell carbon, nitrogen, and chlorophyll a contents, cell volume, photosynthetic carbon assimilation vs. irradiance, short-term uptake of ammonium and nitrate vs. their ambient concentrations, and in vitro activities of the assimilatory enzymes nitrate reductase and glutamic dehydrogenase. The various parameters showed either an increase (pattern a) or a decrease (pattern b) with increasing N limitation. Those following pattern a were nitrate reductase activity and the capacity to assimilate nitrate and ammonium. Those following pattern b were glutamic dehydrogenase activity, photosynthetic rate, nitrogen:carbon and chlorophyll a:carbon composition ratios. Results are discussed in terms of the interpretation such measurement for natural phytoplankton and effects of circadian periodicity.     

Alloimmune interactions of a lymphoproliferative disease-inducer gene Arp and linkage to Pep-7

In this report we describe a new diallelic gene system, Arp ^aand Arp ^b, which codes for surface antigens on murine lymphocytes. Arp ^bis present only in a mutant strain of BALB/c which is designated BALB/c-Arp ^b. Normal BALB/c and all other strains of mice tested express Arp ^a. The Arp ^bmutation is associated with a newly discovered polymorphism of the Peptidase-7 enzyme, Pep-7^b, which codes for a variant form of the enzyme with a faster anodal mobility on electrophoresis than the commonly known form. The Arp locus controls a range of alloimmune interactions between Arp incompatible lymphocytes. These include mixed lymphocyte reactivity, host-versus-graft and graft-versus-host reactions and the development of weak cytotoxic but strong cytostatic effector lymphocytes which are allo- as well as autoreactive. The association between Arp and Pep-7 and the biological significance of the Arp locus are discussed.     

Expression of a chimaeric heat-shock-inducible Agrobacterium 6b oncogene in Nicotiana rustica

The T-6b gene of Agrobacterium tumefaciens strain Tm4 induces tumours on Nicotiana rustica by an as yet unknown mechanism. These tumours cannot be regenerated into normal plants. To study the effect of the T-6b gene product on normal plant cells, the T-6b gene was placed under control of the Drosophila melanogaster hsp70 heat-shock promoter and introduced into N. rustica. Progeny of an hsp70-T-6b transformant developed into normal plants. The inducibility of the hsp70-T-6b construct was shown by northern analysis and by heat-shock-dependent growth alterations on the level of whole seedlings. Upon wounding at normal temperature conditions hsp70-T-6b plants formed small tumours on leaves and stems. Grafts between transformed plants and normal plants led to a wound callus which remained limited to transformed tissues, indicating that the T-6b gene product does not diffuse. Protoplasts of hsp70-T-6b plants divided in the same way as control protoplasts under standard culture conditions. However, when protoplast cultures were started in the absence of hormones, normal cells rapidly lost their sensitivity towards hormones, whereas hsp70-T-6b cells remained sensitive for a significantly longer period. Thus, the T-6b gene product alters hormone sensitivity during the initial phases of protoplast culture.     

Thermal time and ecological strategies - a unifying hypothesis

Rates of embryogenesis and of development and growth in several nematodes are linearly related to temperature over a considerable range. On this basis, published data on the thermal time requirements are compared for a tropical and a temperate species of plant parasitic nematode Meloidogyne javanica and M. hapla respectively, the two being closely related and morphologically and biologically similar. M. hapla has a lower base temperature (T_b) and a higher thermal constant (S) than M. javanica with the relative values being almost inversed. Consequently, above their respective T_b values the slope of the relationship between rate of development and temperature was greater for the tropical species than that for the temperate species. A mathematical exploration of the relationship between T_b and S was made assuming that, over a narrow range, T_b×S was a constant. With this assumption, for any given average environmental temperature (T_e) the optimum base temperature for minimum developmental duration was T_e/2, and the temperature at which the duration of development was equal for the otherwise identical species was shown to be the sum of their base temperatures. The practical effect of the differences in T_b and S was to give M. hapla, the temperate species, a shorter life cycle and hence a competitive advantage at temperatures below 21ÅC and M. javanica, the tropical species, the advantage above that temperature. It is argued that a negative correlation between T_b and S is likely to be widespread, and provides a mechanism for regulating the distributions of related, competing organisms. Support for the hypothesis that the value of S tends to decrease as T_b increases is derived from data on the embryogenesis of an animal parasitic nematode Haemonchus contortus and from seed germination studies. Contrary results and exceptions are also briefly discussed. The observed interaction between T_b and S may be fundamental to many poikilothermic organisms and plants and provides an explanation for tropical species generally having higher T_b values than temperate ones. The ecological implications of different values of T_b and of S, including their relationship with organisms which are “r” or “K” strategists are briefly discussed.     

The Photoxidative Alteration of Chlorophylls in Methyl Linoleate and Prooxidant Activity of Their Decomposition Products

Methyl linoleate containing chlorophylls and/or pheophytins was exposed to light in the presence of oxygen. The photooxidative reaction of both chlorophylls a and b was first-order, and the reaction rate for chlorophyll a was higher than that for chlorophyll b. On the other hand, pheophytins a and b hardly decomposed even after irradiation for 24 hr, and retained a green or a brownish-green color. In qualitative analysis of the photooxidation products of chlorophylls a and b, no pheophytins or pheophorbides were detected, while green and polar red pigments were observed on a thin layer chromatogram near the spot of chlorophyll and the origin, respectively. These photooxidation compounds also had prooxidant effects as well as did chlorophyll.