Complex Nutrient Medium Research Articles

Towards a standardised orchid germination protocol: determining the optimal duration of chemical scarification for orchid seeds by means of viability and permeability staining

Complex nutrient media are used for the germination of orchids in the laboratory. These substrates are also suitable for the growth of microbes and thus the sterilization of both the medium and the seeds prior to sowing is mandatory. The surface disinfection of orchid seeds with chemical substances eliminates possible infections and enhances germination in several cases, by chemically scarifying their seed coats. However, long durations of chemical treatments may result in viability loss, and consequently, precisely determining the optimal treatment duration is essential for developing effective germination protocols. In this study, the double staining technique with Tetrazolium Chloride and Trypan Blue is tested as a proxy method to determine the optimal chemical treatment duration for 10 orchid taxa of Greece. Various durations of chemical treatment with 5% Ca(OCl)2 were carried out on independent seed samples which were subsequently tested, pairwise, for both germination and viability. The results in the corresponding experiments are extraordinarily similar and we conclude that the double viability staining is a quick and accurate technique in determining the permeability and optimal duration for chemical scarification prior to germination experiments. The storability potential of the taxa studied is discussed in comparison with previous data of the NKUA Seed Bank.

MICROBIAL GROWTH OF LACTOBACILLUS DELBRUECKII SSP. BULGARICUS B1 IN A COMPLEX NUTRIENT MEDIUM (MRS-BROTH)

The microbial growth of the probiotic strain Lactobacillus delbrueckii ssp. bulgaricus B1, cultivated in a complex nutrient medium (MRS-broth), was studied in the present work. The complex nutrient medium provides not only the carbon source necessary for the growth of biomass, but also all the additional sources of nitrogen, phosphorus and other components that the biomass needs for its growth. The use of non-structural mathematical dependences determines the optimal conditions (substrate concentration) for the accumulation of biomass or lactic acid, depending on the needs of the specific production.

Biopreservation of Chocolate Mousse with Lactobacillus helveticus 2/20: Microbial Challenge Test

Probiotic bacteria are used for food biopreservation because their metabolic products might contribute to ensuring food microbiological safety and/or increase its shelf life without the addition of chemical preservatives. Moreover, biopreserved foods are excellent vehicles for the delivery of probiotic bacteria. The aim of the study was to investigate the potential of chocolate mousse food matrix for the delivery of the probiotic strain Lactobacillus helveticus 2/20 (Lb. helveticus 2/20) and to investigate its capacity to inhibit the growth of two foodborne pathogenic bacteria (Staphylococcus aureus and Escherichia coli). Therefore, the populations of free or encapsulated in calcium alginate Lb. helveticus 2/20 cells and/or of each pathogen (used to voluntarily contaminate each sample) were monitored both in complex nutrient medium (MRS broth) and in chocolate mousse under refrigeration conditions and at room temperature. Lb. helveticus 2/20 alone in free or encapsulated state effectively inhibited the growth of Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923 in chocolate mousse when stored at 20 ± 2 °C. Practically no viable unwanted bacteria were identified on the 7th day from the beginning of the process. High viable Lb. helveticus 2/20 cell populations were maintained during storage under refrigerated conditions (4 ± 2 °C) and at room temperature. Chocolate mousse is thus a promising food matrix to deliver probiotic Lb. helveticus 2/20 cells, which could also protect it from contamination by unwanted bacteria.

Influence of the composition of nutrient media on the productivity and biological activity of the strain of entomopathogenic fungus Beauveria bassiana

The productivity and biological activity of the entomopathogenic fungus Beauveria bassiana (F-145) strain during the liquid-phase cultivation on various substrates for developing a biopesticide in its native form was analyzed (the research was carried out in 2019). For submerged cultivation, by-products from dairy and beer production (milk whey and brewing spent grain liquor) were used as components of the nutrient medium with addition of diesel fuel (DF) and Tween-80 as inducers of biological activity. It has been established, that the productivity of the strain on industrial by-product substrates was 1.5-2 times higher than on the Czapek medium. A high yield of a mycelial biomass with a titer of 108 -1010 CFU/ml was shown in a 5-day suspension based on a mixture of milk whey and brewing spent grain liquor. The biological activity of the culture suspension of the strain was determined. It was shown that the nematicidal activity of Beauveria bassiana strain with regard to nematodes of the Rhabditis sp. was largely manifested in a suspension obtained on a mixed medium with the addition of inducers. Ninety per cent death at mobile nematode stages was registered within one or two days of test-organism incubation. A complex nutrient medium composition containing by-products and inducers contributed to the preservation of the biological activity of the strain. The strain nematicidal activity was established at the level of 67-80 per cent with a titer of 106 -107 CFU/ml when the suspension was stored for 67 days.

Comparative Analysis of Different Nutrient Media for Growth of Agrobacterium tumefaciens under Small Volume Cultures

Aims: To assess the effects of different selective complex growth media: Yeast Extract Peptone (YEP), Yeast Extract Mannitol (YEM), and Luria Bertani (LB) on growth and multiplication of three different strains of Agrobacterium tumefaciens.
 Place and Duration of Study: Molecular Biology Laboratory, Genetics and Tree Improvement Division, Arid Forest Research Institute, Jodhpur, Rajasthan, India, between July and December 2020.
 Methodology: Three strains of Agrobacterium (EHA105, GV3101 and LBA4404) were inoculated to grow on three different complex nutrient media (YEP, YEM, LB) supplemented with respective antibiotics for a period of 30 hours. Optical Density (O.D) at 600 nm was measured at every 3-hour interval to analyze differential growth pattern.
 Results: Though all the three nutrient media supported the growth of bacteria, YEP medium supported the fast growth of all strains of bacteria, LB medium was a little less efficient but comparable to YEP media. However, YEM medium proved the least supportive for bacterial growth among the three media types.
 Conclusion: Agrobacterium is used for genetic transformation of plant species and crop improvement. knowledge of the growth pattern of bacteria is necessary for the effective infection of plant material. We have analyzed that three strains of Agrobacterium on three different complex media, among the three media YEP media was supporting fast growth. This study can help the researchers to gain knowledge of the growth pattern of Agrobacterium to be used in the genetic transformation of plant species accordingly.

Study of the Effect of Lower Eukaryotes on Tacrolimus (FK-506) Biosynthesis by the Streptomyces tsukubensis Strain VKM Ac-2618D

The effect of inactivated yeast cells, individual components of the yeast cells, and the inactivated biomass of filamentous fungi (micromycetes) of various taxonomic positions on the biosynthesis of immunosuppressant tacrolimus (FK-506) by Streptomyces tsukubensis VKM Ac-2618D cells has been studied. The fractions of soluble and insoluble components of yeast cells positively influenced tacrolimus biosynthesis; the FK-506 concentrations reached 447.1± 22.4 mg/L and 514.3 ± 25.7 mg/L, respectively. The insoluble cell components, including the cell walls and cytoplasmic membranes, had a greater effect than other fractions. The presence of whole baker’s yeast cells and the filamentous fungus Aspergillus ochraceus biomass in the complex nutrient medium stimulated tacrolimus production by S. tsukubensis VKM Ac-2618D (667.5 ± 33.4 mg/L and 633.5 ± 31.7 mg/L, respectively). The biomass of deuteromycete Curvularia lunata had no effect on FK‑506 biosynthesis (288.4 ± 14.4 mg/L). Conversely, the biomass of the Gongronella butleri fungus fully inhibited growth and tacrolimus production in S. tsukubensis. The increase in FK-506 biosynthesis in the presence of the cells of the tested micromycetes is more likely due to the polysaccharide composition of their cell walls than to the ergosterol content. The results expand the knowledge of S. tsukubensis physiology and may be useful for the development of the biotechnological process of tacrolimus production.

Microbiological synthesis optimization of probiotic microorganisms Lactobacillus plantarum using combined nutrient media based on AIC secondary raw materials

The modern agro-industrial complex of the Russian Federation produces a significant number of by-products containing valuable nutrients. The article considers the possibilities of using secondary raw materials of agro-industrial sector in biotechnology industry. The role of waste processing of agricultural raw materials in optimization of lactobacilli cultivation process has been studied. Experimental studies of growth dynamics of probiotic microorganisms Lactobacillus plantarum on complex nutrient media using food and agriculture production waste were carried out. The optimal composition of the nutrient medium for lactobacilli growth and the time of microorganisms cultivation corresponding to the maximum increase of biomass were determined. Comparative analysis of media components for growing Lactobacillus plantarum according to the growth index of probiotic microorganisms was carried out. The growth stimulating role of such substrates for cultivation as beet pulp and corn gluten has been established.

Определение параметров глубинного культивирования микромицета Trichoderma asperellum ВКПМ F-1323 для получения споровых форм культуры

The basic physical parameters of the cultivation of micromycete Trichoderma asperellum VKPM F-1323 are determined on complex nutrient media. The choice of the object of study was due to its antagonistic activity with attitude to many phytopathogenic fungi such as Ascochyta pisi, Cercospora beticola, Claviceps purpurea, Sclerotinia sclerotiorum, Alternaria alternata, Botrytis cinerea, Fusarium graminearum, Passalora fulva, Verticillum dahliae, as well as growth-stimulating activity with attitude to plants, suggesting a motivation for the use of Trichoderma asperellum VKPM F-1323 as the basis for biological products used in agriculture. During the experiment, the nutrient medium composition (g/l) was used: molasses – 20, yeast extract – 7, NaNO3 – 2, K2HPO4 – 1, KCl – 0.5, MgSO4 – 0.5, FeSO4 – 0.01. Before sterilization, the pH was adjusted to 7.5. Cultivation was carried out at a temperature of 27 °C with constant stirring and aeration in autoclaved laboratory fermenters with a working volume of 2 l. The cultivation of micromycete in a liquid nutrient medium was chosen in connection with the rapid progress of the process. In addition, unlike solid-phase cultivation, the deep one allows accumulating in the finished product form numerous secondary metabolites with antagonistic activity against phytopathogens, as well as growth-stimulating activity with attitude to plants. In the work, the values of mixing speed, aeration intensity, and initial pH of the medium are determined, which determine the maximum productivity of the process according to the conidia concentration (titer) of the studied culture. During the cultivation process, the pH level was monitored without maintaining it during the process, since it is known that a change in pH values characterizes the course of the cultivation process with the formation of spore forms, and the initial pH value is an important factor in the conidia formation of Trichoderma fungi. It is shown that during cultivation for 72 hours the maximum concentration of conidia is achieved for a stirring speed of 700 rpm, without reflective baffles (chippers) in the design of the fermenter, aeration intensity of 0.25 l/l of medium×min and the initial pH of the nutrient medium 7.0. Subject to the above conditions, the concentration of conidia is 1.35 ± 0.09 conidia /ml.

Влияние низших грибов на биосинтез такролимуса (FK-506) штаммом Streptomyces tsukubensis BKM Ас-2618Д

The effect of inactivated yeast cells, cell components of yeast cells, and inactivated biomass of the lower fungi of various taxonomic positions on the immunosuppressant tacrolimus (FK-506) biosynthesis by Streptomyces tsukubensis VKM Ac-2618D has been studied. The fractions of soluble and insoluble components of yeast cells positively influenced the tacrolimus biosynthesis with its concentrations reaching 447.1±22.4 mg/L and 514.3±25.7 mg/L, respectively. Wherein, the insoluble cell components including cell walls and cytoplasmic membranes had a greater effect. The presence of the baker's yeast whole cells and filamentous fungus Aspergillus ochraceus biomass in the complex nutrient medium stimulated the tacrolimus production by S. tsukubensis VKM Ac-2618D (667.5±33.4 mg/L and 633.5±31.7 mg/L, respectively). The biomass of deuteromycete Curvularia lunata had no effect on the FK-506 biosynthesis (288.4±14.4 mg/L). Conversely, the biomass of fungus Gongronella butleri fully inhibited the growth and tacrolimus production by S. tsukubensis. The increase in the FK-506 production in media containing cells of the lower fungi is mainly related to the polysaccharide composition of their cell walls rather than the ergosterol content. The results extend the knowledge on the S. tsukubensis physiology and could be useful for the development of the tacrolimus biotechnological production. tacrolimus, biosynthesis, Streptomyces tsukubensis, yeasts, fungi, ergosterol The study was supported by the FASIE program UMNIK 16-12 (Project no. 12081GU/2017).

Production of Biosurfactants By Arthrobacter Sp . N3 , a Hydrocarbon Degrading Bacterium

Different screening methods, such as emulsification capacity and oil spreading assays, hydrocarbon overlay agar and modified drop collapse methods were used to detect biosurfactant production by hydrocarbon degrading Arthrobacter sp N3 strain. It was indicated that oil spreading assay was the most reliable method to detect biosurfactant production. To investigate biosurfactant production, batch cultivation of Arthrobacter sp N3 was carried out in a fermenter with complex nutrient medium supplemented by sunflower oil as a carbon source. The highest oil displacement activity was achieved when Arthrobacter sp N3 strain was cultivated in two stages (with aeration for cell production and without aeration for biosurfactant synthesis). Then, two forms of the biosurfactant (crude preparation and partially purified biosurfactant) were recovered from the culture liquid. Furthermore, the biosurfactant produced by Arthrobacter sp N3 strain was analyzed by thin layer chromatography and it was estimated that even a few compounds have surface activity. The effect of temperature and pH on biosurfactant activity was also studied. It was observed that no appreciable changes in biosurfactant activity occurred at temperature and pH values ranges of 4–125 ºC and 5–10, respectively.

Biodegradation of 1,4-dioxane: Effects of enzyme inducers and trichloroethylene

1,4-Dioxane is a groundwater contaminant and probable human carcinogen. In this study, two well-studied degradative bacteria Mycobacterium vaccae JOB5 and Rhodococcus jostii RHA1 were examined for their 1,4-dioxane degradation ability in the presence and absence of its co-contaminant, trichloroethylene (TCE), under different oxygenase-expression conditions. These two strains were precultured with R2A broth (complex nutrient medium) before supplementation with propane or 1-butanol to induce the expression of different oxygenases. Both propane- and 1-butanol-induced JOB5 and RHA1 were able to degrade 1,4-dioxane, TCE, and mixtures of 1,4-dioxane/TCE. Complete degradation of 1,4-dioxane/TCE mixture was observed only in propane-induced strain JOB5. Inhibition was observed between 1,4-dioxane and TCE for all cells. Furthermore, product toxicity caused incomplete degradation of 1,4-dioxane by 1-butanol-induced JOB5. In general, the more TCE degraded, the greater extent of product toxicity cells experienced; however, susceptibility to product toxicity was found to be both strain- and inducer-dependent. The findings of this study provide fundamental basis for developing an effective in-situ remediation method for 1,4-dioxane-contaminated ground water and the first known study of 1,4-dioxane degradation by wild-type strain RHA1.

Cadmium Tolerance by Haloarchaeal Strains Isolated from Solar Salterns of Goa, India

 Abstract—Haloarchaeal strains from solar salterns of Ribandar and Siridao, Goa, India were tested for tolerance to cadmium. The four strains selected named as BBK2, BK6, BS2 and BS17 belonged to different genera of halophilic archaea. An initial screening for various heavy metals [Zn, Cu, Ag, Co and Cd] was performed. Cadmium (Cd) concentrations of 0.5, 1, 2 and 4 mM Cd in complex and glucose synthetic media containing high sodium chloride concentrations were used. Growth was monitored by monitoring optical density (O.D.) values for each of the sample cultures in 24 hr intervals for an incubation period of 5-7 days. Pigmentation and growth was observed in test cultures in complex nutrient media containing cadmium. Interesting results for pigmentation scans of the tested strains having varying peak intensities and exhibiting spectrophotometric profiles associated with typical haloarchaeal pigments were obtained. Correlation between peak intensities of the tested strains and the amount of cadmium in test media was also observed. This investigation shows cadmium tolerance in haloarcheal strains.

“Isolation and Biochemical Characterization of Antibiotic Producing Microorganism from Waste Soil Samples of Certain Industrial areas of India”

The search for new antibiotics continues in a rather overlooked hunting ground. In the course of screening for new antibiotic-producing microorganisms, isolates showing antimicrobial activity were isolated from waste soil samples from various habitats in the Industrial Areas in Dheradun, Uttarakhand, India. Existing methods of screening for antibiotic producers together with some novel procedures were reviewed. Both modified agar-streak and agar-plug methods were used in the primary screens. The use of selective isolation media, with or without antibiotic incorporation and/or heat pretreatment, enhanced the development of certain actinomycete colonies on the isolation plates. Antibiotics have long been considered the magic bullet that would end infectious disease. Although they have improved the health of countless numbers of humans and animals, many antibiotics have also been losing their effectiveness since the beginning of the antibiotic era. Bacteria have adapted defenses against these antibiotics and continue to develop new resistances, even as we develop new antibiotics. In recent years, much attention has been given to the increase in antibiotic resistance. As more microbial species and strains become resistant, many diseases have become difficult to treat, a phenomenon frequently ascribed to both indiscriminate and inappropriate use of antibiotics in human medicine. However, the use of antibiotics and antimicrobials in raising food animals has also contributed significantly to the pool of antibiotic resistant organisms globally and antibiotic resistant bacteria are now found in large numbers in virtually every ecosystem on earth. Dual culture bioassays were used to screen seven selected Bacillus isolates for activity against four plant pathogenic fungi in vitro. All isolates were able to inhibit the pathogens to varying degrees. Two isolates, R29 and B81, were selected for further testing and characterization. Further bioassays were performed on five complex nutrient media which were adjusted to pH S.S and 7, and both incubated at 2SoC and 30°C respectively. It was found that pH and media composition showed significant influences on the antifungal activities of the isolates tested, but that a SoC temperature difference in incubation temperature did not. Tryptone soy agar was found to give rise to the largest inhibition zones. Both isolates were tentatively identified using standard biochemical and morphological tests. Based on its phenotypic characteristics, R29 was identified as a strain of B. subtilis. B81 proved to be more difficult to assign to a specific group or species of Bacillus, though B. subtilis and B. licheniformis were considered to be the nearest candidates. Genomic DNA was extracted from both isolates and a portion of each of their 16s rDNA genes were amplified and sequenced for homology testing against the GeneBank database. Homology testing confirmed that both isolates were members of the genus Bacillus and most probably strains of B. subtilis. The DNA fragment used for sequencing proved to be too small to give conclusive identification of the isolates.

Isolation and characterization of a facultative methanotroph degrading malodor-causing volatile sulfur compounds

Simultaneous removal of methane and malodor-causing volatile sulfur compounds (MVSCs), both emitted from landfills, is a desirable characteristic for methane-mitigation approaches. A methanotrophic bacterium was isolated from a microbial consortium, enriched with methane and dimethyl sulfide (DMS). It grew in the complex nutrient medium R2A without methane, and stably exhibited methanotrophic activity after facultative growth. It was identified as Sphingopyxis sp. MD2 by comparison of the 16S rRNA gene. It belongs to Sphingomonadales, whose members have not shown methanotrophic activity, phylogenetically distinct from orders of known methanotrophs. The MD2 biomass increased at a growth rate of 1.18d−1 when methane was used as the sole growth substrate. An inhibition test with allylthiourea and PCR/sequencing confirmed the presence of particulate methane monooxygenase in MD2. DMS decreased the methane oxidation rate (2634±146μmolegDCW−1h−1) by 12%, while H2S had no effect on the methane oxidation rate. Interestingly, methanethiol (MT) enhanced the methane oxidation rate by more than 50%. MD2 degraded H2S and MT, regardless of the presence of methane. MD2 also degraded DMS in the presence of methane, indicating co-metabolism. These combined results indicate that MD2 may be a promising biological resource for simultaneous removal of methane and MVSCs.

Biodegradation of triclosan by a wastewater microorganism

Triclosan, a synthetic antimicrobial agent, has been considered as an emerging environmental contaminant. Here we reported a triclosan-degrading wastewater bacterial isolate, Sphingopyxis strain KCY1, capable of dechlorinating triclosan with a stoichiometric release of chloride. The stain can degrade diphenyl ether but not 2,4,4′-tribromodiphenyl ether and 2,2′,4,4′-tetrabromodiphenyl ether, despite all these three compounds are structurally similar to triclosan. While strain KCY1 was unable to grow on triclosan and catechol, it could grow with glucose, sodium succinate, sodium acetate, and phenol. When grown with complex nutrient medium containing a trace amount of triclosan (as low as 5 μg/L), the strain could retain its degradation ability toward triclosan. The maximum-specific triclosan degradation rate (qm) and the half-velocity constant (Km) are 0.13 mg-triclosan/mg-protein/day and 2.8 mg-triclosan/L, respectively. As triclosan degradation progressed, five metabolites were identified and these metabolites continue to transform into non-chlorinated end products, which was supported by a sharp drop in androgenic potential. The activity of catechol 2,3-dioxygenase in the cell extract was detected. No triclosan degradation was observed in the presence of 3-fluorocatechol, an inhibitor of meta-cleavage enzyme, suggesting that triclosan degradation proceed via meta-cleavage pathway. Based on all the observations, a degradation pathway for triclosan by strain KCY1 was proposed.

Advanced Stoichiometric Analysis of Metabolic Networks of Mammalian Systems

Metabolic engineering tools have been widely applied to living organisms to gain a comprehensive understanding about cellular networks and to improve cellular properties. Metabolic flux analysis (MFA), flux balance analysis (FBA), and metabolic pathway analysis (MPA) are among the most popular tools in stoichiometric network analysis. Although application of these tools into well-known microbial systems is extensive in the literature, various barriers prevent them from being utilized in mammalian cells. Limited experimental data, complex regulatory mechanisms, and the requirement of more complex nutrient media are some major obstacles in mammalian cell systems. However, mammalian cells have been used to produce therapeutic proteins, to characterize disease states or related abnormal metabolic conditions, and to analyze the toxicological effects of some medicinally important drugs. Therefore, there is a growing need for extending metabolic engineering principles to mammalian cells in order to understand their underlying metabolic functions. In this review article, advanced metabolic engineering tools developed for stoichiometric analysis including MFA, FBA, and MPA are described. Applications of these tools in mammalian cells are discussed in detail, and the challenges and opportunities are highlighted.

Host cell-free growth of the Q fever bacterium Coxiella burnetii

The inability to propagate obligate intracellular pathogens under axenic (host cell-free) culture conditions imposes severe experimental constraints that have negatively impacted progress in understanding pathogen virulence and disease mechanisms. Coxiella burnetii, the causative agent of human Q (Query) fever, is an obligate intracellular bacterial pathogen that replicates exclusively in an acidified, lysosome-like vacuole. To define conditions that support C. burnetii growth, we systematically evaluated the organism's metabolic requirements using expression microarrays, genomic reconstruction, and metabolite typing. This led to development of a complex nutrient medium that supported substantial growth (approximately 3 log(10)) of C. burnetii in a 2.5% oxygen environment. Importantly, axenically grown C. burnetii were highly infectious for Vero cells and exhibited developmental forms characteristic of in vivo grown organisms. Axenic cultivation of C. burnetii will facilitate studies of the organism's pathogenesis and genetics and aid development of Q fever preventatives such as an effective subunit vaccine. Furthermore, the systematic approach used here may be broadly applicable to development of axenic media that support growth of other medically important obligate intracellular pathogens.

A Soy-Based Alternative to Traditional Bacterial Nutrient Media

A vegan is an individual who does not use or consume products of any kind (Vegan Society, accessed June 15, 2008). The term animal product in a vegan context refers to any material derived from animals for human use (Stepaniak, 2000), such as meat, poultry, seafood, eggs, dairy products, honey, fur, leather, wool, and silk (Vegan Society, accessed June 15, 2008). Common by-products include gelatin, lanolin, whey, casein, and beeswax (Vegan Society, accessed June 15, 2008). Reasons cited for choosing a vegan/vegetarian lifestyle include ethical or moral convictions concerning rights, concerns for the environment, human health, and religious concerns (Time/CNN, July 7, 2002; Social Survey Division of the Office for National Statistics, 2002; Stepaniak, 2000). Various polls have reported that vegans make up anywhere from 0.2% to 1.4% of the adult U.S. population, a group that seems to be growing (Time/CNN, July 7, 2002; Stahler, 2006). Further, most people in that group fall between the ages of 16 and 34 (The Vegan Research panel, accessed June 15, 2008, an age range typical of most college students. Most complex nutrient media used in a general microbiology lab (Nutrient Broth, Luria-Bertani Broth, or Trypticase Soy Broth) contain animal-derived ingredients, such as casein, tryptone, or meat extracts. Our goal was to produce a medium that is free of all products, yet supports the growth of most non-fastidious bacteria in a manner comparable to commercially available animal-based media. To provide the growing vegan population (and anyone else uncomfortable with using products) a viable alternative in microbiology lab, we wanted to develop an equally-effective, inexpensive, animal-free substitute to conventional media. We initially tested several combinations of soy flour and other additives on eight species of bacteria, and qualitatively compared the amount of growth on these plates against standard animal-based nutrient agar plates. From these trials, we found that a mixture of toasted soy flour (SoyQuik[R]) and yeast extract powder yielded the best, most cost-effective results. Quantitative and qualitative analyses were performed to compare growth of bacterial cultures in our soy-based media to growth in the standard animal-based media. As a final assessment, we tested the soy-based medium side-by-side against animal-based media typically recommended by experimental protocol during standard microbiology labs. * Methods Nutrient Media Preparation Prospective animal-free additives were purchased in health food sections of local grocery stores or ordered online at the EnerGFoods Web site (Ener-G.com). These products (including toasted soy flour, potato flour, and rice bran) were incorporated in varying ratios into agar plates. Commercially-available Yeast Extract and Nutrient Broth (Difco[R]) powders were also used and prepared according to the manufacturer's instructions. All ingredients were dissolved in distilled water and sterilized as is typical in nutrient media preparation. When solid media was required, the preparation included 1.5% agar. Media recipes and their corresponding designations include: * SF = 3% soy flour * SP = 3% soy flour + 2% potato flour * SR = 3% soy flour + 2% rice bran * SY = 3% soy flour + 2% yeast extract * SPY = 3% soy flour + 2% potato flour + 2% yeast extract * SRY = 3% soy flour + 2% rice bran + 2% yeast extract * NB = nutrient broth (made according to manufacturer's instructions) * NBY = nutrient broth + 2% yeast extract * YE = 2% yeast extract * 2SF = 2% soy flour * MSY (modified soy media) = 2% soy flour + 0.5% yeast extract Qualitative Growth Analyses Fresh, single colonies (

Growth of the Thermophilic Bacterium Geobacillus uralicus as a Function of Temperature and pH: An SCM-Based Kinetic Analysis

The synthetic chemostat model (SCM), originally developed to describe nonstationary growth under widely varying concentrations of the limiting substrate, was modified to account for the effects of nontrophic factors such as temperature and pH. The bacterium Geobacillus uralicus, isolated from an ultradeep well (4680 m), was grown at temperatures ranging from 40 to 75°C and at pH varying from 5 to 9. The biomass kinetics was reasonably well described by the SCM, including the phase of growth deceleration observed in the first hours after a change in the cultivation temperature. At an early stage of batch growth in a neutral or alkalescent medium, bacterial cells showed reversible attachment to the glass surface of the fermentation vessel. The temperature dependence of the maximum specific growth rate (μm) was fitted using the equation μm = A exp (λT )/{1 + expB[1 – C/(T + 273)]}, where A, λ, B, and C are constants. The maximum specific growth rate of 2.7 h–1 (generation time, 15.4 min) was attained on a complex nutrient medium (peptone and yeast extract) at 66.5°C and pH 7.5. On a synthetic mineral medium with glucose, the specific growth rate declined to 1.2 h–1, and the optimal temperature for growth decreased to 62.3°C.

Demonstration of residual antibacterial activity in plasma of vaccinated Penaeus vannamei

Plasma from naı̈ve Penaeus vannamei juveniles lacked antibacterial factors and significantly enhanced the growth of Escherichia coli strain XL1-Blue MRF′ compared with sea water complex (SWC) nutrient medium. In contrast, plasma from prawns previously injected with both live, non pathogenic Vibrio harveyi strain DPEX and formalin-killed cells of V. harveyi strains BP04, DPEX, E. coli and sterile shrimp salt solution (SSS), inhibited bacterial growth. Antibacterial activity appeared within 6 h and was detectable until 7 days after “vaccine” administration. Lysozyme activity was not detected in plasma from vaccinated prawns. Both in vitro and in vivo bacterial killing were facilitated by vaccine administration. Activation of the prophenoloxidase (proPO) system was independent of bacterial virulence.