Process optimization and modeling for the cultivation of Nannochloropsis sp. and Tetraselmis striata via response surface methodology

The aim of this study was to determine the optimal physical process conditions for the cultivation of locally isolated strains of Nannochloropsis sp. and Tetraselmis striata to achieve maximum growth rate. It was essential to evaluate biomass production at different agitation rates, light intensities, and temperature levels. Central composite design and response surface methodology were applied to design the experiments and optimize the cultivation process for Nannochloropsis sp. and T. striata. The specific growth rate of 0.250 d^?1 was obtained for Nannochloropsis sp. cells under the light intensity of 54 μmol photons · m^?2 · s^?1, at the agitation rate of 151 rpm in 24.5°C. The optimal physical process conditions for T. striata were obtained under the light intensity of 56 μmol photons · m^?2 · s^?1 in 25.5°C at the agitation rate of 151 rpm in 25.5°C, resulting in a specific growth rate of 0.226 d^?1. The predicted values were justified by the verification tests. Good agreement between the predicted values and the experimental values confirmed the validity of the models for the cultivation of microalgal strains. In this article, the noteworthy result was that temperature was a dominant factor in obtaining high chl‐a content for Nannochloropsis sp., whereas the growth of T. striata strongly depended on light exposure.     

Germanium toxicity in selected bacterial and yeast strains

Summary The toxicity of germanium dioxide (GeO₂) to 21 bacterial and 13 yeast strains was investigated in liquid broth medium to obtain information on strains tolerant to high (1 to 2 mg/ml) GeO₂ concentrations.Arthrobacter sp. NRC 32005,enterobacter aerogenes NRC 2926,Klebsiella aerogenes NCTC 418 andPseudomonas putida NRC 5019 were tolerant to 1 mg/ml GeO₂.Bacillus sp. RC607 was able to grow in the presence of 2 mg/ml GeO₂ at pH 10 in broth culture. The yeastsCandida guilliermondii, Candida shehatae andPachysolen tannophilus were the most sensitive to GeO₂ as evidenced by their diminished growth rates at a GeO₂ concentration as low as 0.1 mg/ml. None of the yeast strains tested exhibited growth in the presence of 1 mg/ml GeO₂. The high pH of the medium containing germanium may be partially responsible for the growth inhibition of the yeast cultures. Select bacterial cultures previously exposed to 1 mg/ml GeO₂ could tolerate and grow better at 2 mg/ml GeO₂, suggesting the existence of very efficient adaptive mechanisms. The pH of the medium could modulate GeO₂ tolerance and this effect was found to be strain-dependent.     

Evaluation of halophilic microalgae isolated from Rabigh Red Sea coastal area for biodiesel production: Screening and biochemical studies

In the present study, different water samples from Red Sea coastal area at Rabigh city, Saudi Arabia were studied for their dominant algal species. Microalgal isolation was carried out based on dilution method and morphologically examined using F/2 as a growth medium. Dry weight and main biochemical composition (protein, carbohydrates, lipids) of all species were performed at the end of the growth, and biodiesel characteristics were estimated. Nannochloropsis sp., Dunaliella sp., Tetraselmis sp., Prorocentrum sp., Chlorella sp., Nitzschia sp., Coscinodiscus sp., and Navicula sp. were the most dominant species in the collected water samples and were used for further evaluation. Nannochloropsis sp. surpassed all other isolates in concern of biomass production with the maximum recorded dry weight of 0.89 g L^?1, followed by Dunaliella sp. (0.69 g L^?1). The highest crude protein content was observed in Nitzschia sp. (38.21%) and Dunaliella sp. (18.01%), while Nannochloropsis sp. showed 13.38%, with the lowest recorded lipid content in Coscinodiscus sp. (10.09%). Based on the growth, lipid content, and biodiesel characteristics, the present study suggested Dunaliella sp. and Nitzschia sp. as promising candidates for further large-scale biodiesel production.     

Growth of Particle-Bearing and Particle-Free Paramecium aurelia in Axenic Culture*

SYNOPSIS. Several strains of particle-bearing and particle-free Paramecium aurelia have been cultivated in an axenic medium composed of proteose peptone, trypticase, yeast nucleic acid, MgSO⁴.7H²O, TEM-4T (diacetyl tartaric acid esters of tallow monoglycerides), stigmasterol and a mixture of vitamins. The “yeast fraction,” an indispensable component of previous media used for the cultivation of these ciliates has been replaced by a mixture of trypticase, yeast nucleic acid and TEM-4T. Particle-bearing animals of stock 299 lambda, 138 mu, and 139 pi maintain their particles when cultivated in the medium, whereas particle-bearing animals of stock 51 kappa, 225 kappa and 114 signia do not. With the exception of stock 92 (syngen 3) the medium appears to be selective in its ability to support the growth of animals of the even- but not odd-numbered syngens of P. aurelia. Maintenance of the particles was dependent only to a small degree upon environmental conditions brought about by changes in pH and temperature. Division of the particles was found to be comparable with the division of the protozoan. Methods for the growth, maintenance and mass cultivation of particle-bearing P. aurelia are given in detail.     

Genomic-assisted characterisation of Pseudomonas sp. strain Pf4, a potential biocontrol agent in hydroponics

In an attempt to select potential biocontrol agents against Pythium spp. and Rhizoctonia spp. root pathogens for use in soilless systems, 12 promising bacteria were selected for further investigations. Sequence analysis of the 16S rRNA gene revealed that three strains belonged to the genus Enterobacter, whereas nine strains belonged to the genus Pseudomonas. In in vitro assays, one strain of Pseudomonas sp., Pf4, closely related to Pseudomonas protegens (formerly Pseudomonas fluorescens), showed noteworthy antagonistic activity against two strains of Pythium aphanidermatum and two strains of Rhizoctonia solani AG 1-IB, with average inhibition of mycelial growth >80%. Strain Pf4 was used for in vivo treatments on lamb’s lettuce against R. solani root rot in small-scale hydroponics. Pf4-treated and untreated plants were daily monitored for symptom development and after two weeks of infection, a significant protective effect of Pf4 against root rot was recorded. The survival and population density of Pf4 on roots were also checked, demonstrating a density above the threshold value of 10⁵?CFU?g^?1 of root required for disease suppression. Known loci for the synthesis of antifungal metabolites, detected using PCR, and draft-genome sequencing of Pf4 demonstrated that Pseudomonas sp. Pf4 has the potential to produce an arsenal of secondary metabolites (plt, phl, ofa and fit-rzx gene clusters) very similar to that of the well-known biocontrol P. protegens strain Pf-5.     

In vitro degradation of wheat straw by anaerobic fungi from small ruminants

Abstract

Anaerobic ruminal fungi may play an active role in fibre degradation as evidenced by the production of different fibrolytic enzymes in culture filtrate. In the present study, 16 anaerobic fungal strains were isolated from ruminal and faecal samples of sheep and goats. Based on their morphological characteristics they were identified as species of Anaeromyces, Orpinomyces, Piromyces and Neocallimastix. Isolated Neocallimastix sp. from goat rumen showed a maximum activity of CMCase (47.9 mIU ml^?1) and filter paper cellulase (48.3 mIU ml^?1), while Anaeromyces sp. from sheep rumen showed a maximum xylanolytic activity (48.3 mIU ml^?1). The cellobiase activity for all the isolates ranged from 178.0 – 182.7 mIU ml^?1. Based on the enzymatic activities, isolated Anaeromyces sp. from sheep rumen and Neocallimastix sp. from goat rumen were selected for their potential of in vitro fibre degradation. The highest in vitro digestibility of NDF (23.2%) and DM (34.4%) was shown for Neocallimastix sp. from goat rumen, as compared to the digestibility of NDF and DM in the control group of 17.5 and 25.0%, respectively.     

Effect of temperature on growth parameters of psychrophilic bacteria and yeasts

Three bacterial (Pedobacter heparinus, Pedobacter piscium, Pedobacter cryoconitis) and three yeast strains (Saccharomyces cerevisiae, Leucosporidiella creatinivora, Rhodotorula glacialis) of different thermal classes (mesophiles and psychrophiles) were tested for the effect of temperature on a range of growth parameters, including optical density, viable cell numbers, and cell dry mass, in order to determine the temperature conditions under which maximum biomass formation is obtained. Maximum values of growth parameters obtained at the stationary growth phase of the strains were used for statistical calculation. Temperature had a significant (P ≤ 0.05) effect on all growth parameters for each strain; correlations between the growth parameters were significant (P ≤ 0.05–0.01). The maximum growth temperature or the temperature at which microbial growth was fastest was in no case the temperature at which the investigated strains produced the highest amount of biomass. All tested psychrophilic bacteria and yeast strains produced highest amounts of cells (as calculated per mg cell dry mass or per OD₆₀₀ unit) at 1°C, while cell numbers of mesophiles were highest at 20°C. Thus, cultivation temperatures close to the maximum growth temperature are not appropriate for studying psychrophiles.     

<Emphasis Type="Italic">Kazachstania intestinalis</Emphasis> sp. nov., an ascosporogenous yeast from the gut of passalid beetle <Emphasis Type="Italic">Odontotaenius disjunctus</Emphasis>

Three ascosporogenous yeast strains were isolated from the gut of the passalid beetle, Odontotaenius disjunctus, inhabiting on rotten oak trees. DNA sequence comparison and other taxonomic characteristics identified the strains as a novel species in the genus Kazachstania. The name Kazachstania intestinalis sp. nov. (type strain EH085^T = ATCC MYA-4658^T = CBS 11839^T) is proposed for the strains. The yeast is homothallic, producing persistent asci with 1–4 spheroidal ascospores. Molecular phylogeny from ribosomal RNA gene sequences placed this novel species on the basal lineage of a clade including Kazachstania lodderae, Kazachstania exigua, Kazachstania martiniae, and other related Kazachstania spp., but none of those species was a close sister to K. intestinalis.     

A method for the detection of Issatchenkia orientalis in a baker's yeast factory

Summary A method for the detection of Issatchenkia orientalis (Candida krusei), a contaminant of bakers' yeast, is described. Nine different liquid medium types were compared and maximum specific growth rates for the yeast contaminant were determined in each medium. Issatchenkia orientalis grew fastest in malt extract broth (0,37 h^-1) and potato dextrose broth (0,36 h^-1). Five antibiotics were tested for selective inhibition of seven different bakers' yeast strains in malt extract broth. Nystatin was the only antibiotic tested that inhibited the growth of all the bakers' yeast strains used, but did not affect the growth of I. orientalis.     

Growth model for raceway pond cultivation of Desmodesmus sp. MCC34 isolated from a local water body

Biofuel production by microalgae has the advantage of higher biomass productivity over land crops. The selection of potential microalgae depends on the growth in outdoor mass cultivation during different seasons, which can be predicted by a mathematical model. Here, freshwater green algae were isolated from a local water body in Pilani, Rajasthan, India (geographical coordinates: 28°22′N 75°36′E) and characterized by microscopy and ribosomal RNA analysis. The strain was submitted to the Indian Agricultural Research Institute's microbial culture collection (IARI, India) and identified as Desmodesmus sp. MCC34. This strain, along with a fresh water green algae (Chlorella minutissima), two marine green algae species (Dunaliella salina and Dunaliella tertiolecta) and two nitrogen fixing cyanobacteria (Nostoc muscorum and Anabaena doliolum), were screened for lipid productivity and growth kinetics under culture room and raceway pond conditions. Desmodesmus sp. MCC34 showed the highest specific growth rate (0.26 day^?1), biomass production (1.9 g L^?1) and lipid productivity (103 mg L^?1 day^?1). The optimal temperature and saturating light intensity for maximal growth of Desmodesmus sp. MCC34 were 35 °C and 75 μmol m^?2 s^?1 with molar extinction coefficient of 0.22 m² g^?1, respectively. Desmodesmus sp. MCC34 was then subjected to outdoor cultivation in a 20‐m long raceway pond for 18 days during March and November 2013. The areal biomass productivity and volumetric biomass productivity were 13946.23 kg ha^?1 year^?1 and 56.94 mg L^? 1day^?1 during the month of March, decreasing to 6262.28 kg ha^?1 year^?1 and 25.57 mg L^? 1day^?1 during the month of November. A mathematical model was constructed to explain the relationship between biomass production and growth parameters such as temperature, light intensity and nutrient concentration. The productivity values predicted with the proposed model correspond well with the experimental data, suggesting the validity of the model.     

Growth promotion of maize and lettuce by phosphate-solubilizing Rhizobium leguminosarum biovar. phaseoli

Rhizobium leguminosarum bv. phaseoli strains P31 and R1, Serratia sp. strain 22b, Pseudomonas sp. strain 24 and Rhizopus sp. strain 68 were examined for their plant growth-promoting potential on lettuce and forage maize. All these phosphate solubilizing microorganisms (PSM) were isolated from Québec soils. The plants were grown in field conditions in three sites having high to low amounts of available P. In site 1 (very fertile soil), strains R1 and 22b tended to increase the dry matter yield of lettuce shoots (p≤0.10). Lettuce inoculated with rhizobia R1 had a 6% higher P concentration (p≤0.10) than the uninoculated control. In site 2 (poorly fertile soil), the dry matter of lettuce shoots was significantly increased (p≤0.05) by inoculation with strain P31 and 24 plus 35 kg ha^-1 P-superphosphate, or with strain 68 plus 70 kg ha^-1 P-superphosphate. In site 3 (moderately fertile soil), the dry matter of maize shoots was significantly increased (p≤0.05) by inoculation with strain 24 plus 17.5 kg ha^-1 P-superphosphate, or with strain P31 plus 35 kg ha^-1 P-superphosphate. Inoculation with PSM did not affect lettuce P uptake in the less fertile soil in site 2. In site 3 with the moderately fertile soil, maize plants inoculated with strain R1 had 8% higher P concentration than the uninoculated control (p≤0.01), and 6% with strains P31 and 68 (p≤0.05). The results clearly demonstrate that rhizobia specifically selected for P solubilization function as plant growth promoting rhizobacteria with the nonlegumes lettuce and maize. The P solubilization effect seems to be the most important mechanism of plant growth promotion in moderately fertile and very fertile soils when P uptake was increased with rhizobia and other PSM.     

Activities of the enzymes of formaldehyde catabolism in recombinant strains of <Emphasis Type="Italic">Hansenula polymorpha</Emphasis>

Activities of the enzymes of formaldehyde (FA) catabolism in recombinant strains of the methylotrophic yeast Hansenula polymorpha overproducing NAD⁺- and glutathione-dependent formaldehyde dehydrogenase (FADH) were studied under different cultivation conditions and at elevated FA content. Southern dot-blot analysis confirmed the presence of six to eight copies of the target FLD1 gene in stable recombinant clones of H. polymorpha. Under certain cultivation conditions, the transformants resistant to elevated FA concentrations were shown to produce FADH and other bioanalytically important enzymes: formate dehydrogenase, alcohol dehydrogenase, alcohol oxidase, and formaldehyde reductase. The optimal cultivation conditions for recombinants were determined, resulting in maximum synthesis of FADH: methanol as a carbon source, methylamine as a nitrogen source, FA as an inducer, temperature of 37°C, and cells in the early exponential phase of growth.     

Optimization of cultural conditions for the production of antifungal chitinase by Streptomyces sporovirgulis

Actinomycetes were screened from soil in the centre of Poland on chitin medium. Amongst 30 isolated strains one with high activity of chitinase was selected. It was identified as Streptomyces sporovirgulis. Chitinase activity was detected from the second day of cultivation, then increased gradually and reached maximum after 4 days. The maximum chitinase production was observed at pH 8.0 and 25–30°C in the medium with sodium caseinate and asparagine as carbon and nitrogen sources and with shrimp shell waste as inducer of enzyme. Chitinase of S. sporovirgulis was purified from a culture medium by fractionation with ammonium sulphate as well as by chitin affinity chromatography. The molecular weight of the enzyme was 27 kDa. The optimum temperature and pH for the chitinase were 40°C and pH 8.0. The enzyme activity was characterised by high stability at the temperatures between 35 and 40°C after 240 min of preincubation. The activity of the enzyme was strongly inhibited in the presence of Pb²⁺, Hg²⁺ and stabilized by the ions Mg²⁺. Purified chitinase from S. sporovirgulis inhibited growth of fungal phytopathogen Alternaria alternata. Additionally, the crude chitinase inhibited the growth of potential phytopathogens such as Penicillium purpurogenum and Penillium sp.     

Effect of amaranth on the growth of Rhizobium and Bradyrhizobium strains

The growth rate of different strains of Bradyrhizobium and Rhizobium was studied in media containing amaranth seed meal instead of yeast extract. Results obtained in erlenmeyer flasks and stirred fermenters show that both Bradyrhizobium japonicum strains E109, E110, 5019, 587 and Rhizobium melilotistrains B36, B323, B399, Lq₂₂, Lq₄₂, Lq₅₁ and U₃₂₂, grow satisfactorily in amaranth seed meal medium. Cell count obtained for the strains tested was greater than 4 × 10¹⁰ viable cells.ml^–1. Amaranth seed meal (4 g.l^–1) is a suitable component for culture media that can be used instead of yeast extract.     

Continuous production of diatom Entomoneis sp. in mechanically stirred tank and flat-panel airlift photobioreactors

Continuous production of diatom Entomonies sp. was performed in mechanically stirred tank and flat-panel airlift photobioreactors (FPAP). The maximum specific growth rate of diatom from the batch experiment was 0.98 d^?1. A series of dilution rate and macronutrient concentration adjustments were performed in a stirred tank photobioreactor and found that the dilution rate ranged from 0.7 to 0.8 d^?1 and modified F/2 growth media containing nitrate at 3.09?mg N/L, phosphate at 2.24?mg P/L, and silicate at 11.91?mg Si/L yielded the maximum cell number density. Finally, the continuous cultivation of Entomonies sp. was conducted in FPAP using the optimal conditions determined earlier, resulting in the maximum cell number density of 19.69?×?10⁴ cells/mL, which was approximately 47 and 73% increase from the result using the stirred tank photobioreactor fed with modified and standard F/2 growth media, respectively.     

Synechococcus sp. (PTCC 6021) cultivation under different light irradiances—Modeling of growth rate-light response

Synechococcus sp. (PTCC 6021), a cyanobacterium species, was cultivated in an internally illuminated photobioreactor. The reactor was designed to achieve a monoseptic cultivation of the species. The goal was to study the growth–irradiance behavior of Synechococcus sp. (PTCC 6021). To accomplish this, different initial light irradiances were implemented inside the photobioreactor and the growth of the cells was monitored. It was observed that cell growth increased with higher light intensity until the photoinhibition occurrence at light irradiance higher than 250?μE?m^?2?s^?1. The maximum OD₆₀₀, maximum growth rate, and biomass productivity increased, and hence the extinction coefficient decreased, with the increase in light irradiance before photoinhibition. The maximum optical density (OD₆₀₀) of 5.91 was obtained with irradiance below 250?μE?m^?2?s^?1 during a growth period of 80 days. The modified Monod function could model the growth–irradiance of cells with satisfactory agreement with the experimental data. The comparison of growth–irradiance of the studied species with other photosynthetic organisms showed the same trend as for cyanobacteria with photoinhibition.     

Microbotryozyma collariae gen. nov., sp. nov., a basidiomycetous yeast isolated from a plant bug Collaria oleosa (Miridae)

Two strains of a basidiomycetous yeast were derived from an insect trypanosomatid culture isolated from the intestine of a plant bug, Collaria oleosa (Heteroptera: Miridae), collected in Costa Rica. The yeast did not form ballistoconidia but reproduced only by budding. Teliospores were not observed in individual and crossed cultures of each strain. Morphological and other taxonomic characteristics of the yeast were similar to those of the species in the polyphyletic genus Rhodotorula. However, molecular phylogeny inferred from the internal transcribed spacers and D1/D2 region of the large subunit rRNA gene showed that the strains represent a new species placed among the smut fungi in the family Ustilentylomataceae, which includes Aurantiosporium subnitens, Fulvisporium restifaciens, Ustilentyloma fluitans, and Rhodotorula hordea. Given the well distinguished phylogenetic position of this novel species within the Ustilentylomataceae, we propose Microbotryozyma collariae gen. nov., sp. nov. to accommodate the yeast isolated from C. oleosa, with strain American Type Culture Collection MYA-4666^T (= PRA303-1S = CBS 12537) designated as the type strain.     

Sludge and lignin as growth media for the production of antimicrobial substances by selected microorganisms

Abstract

The ability of sludge, lignin and humic acid to support the growth and production of antimicrobial substances by 19 microorganism strains was tested. Organisms from seven genera: Bacillus, Pseudomonas, Streptomyces, Beauveria, Paecilomyces, Trichoderma and Trichosporon grew on the tested media. On these media, they were also examined for the production of anti-phytopathogenic activity against Fusarium oxysporum P1 BIM, Botrytis cinerea B1 BIM and Pseudomonas syringae. pv. glicinea B-280 BIM. Strains Pseudomonas aurantiaca S-1, Bacillus sp. 19 and Streptomyces sp. 11 synthesize antimicrobial metabolites on the different media. Protease, cellulase, α-amylase, xylanase and peroxidase were also produced by the same microorganisms, which can explain their ability to grow on sludge, and laccase was produced only by P. aurantiaca S-1. HPLC analysis of the lignin culture media before and after P. aurantiaca S-1 growth has shown that the major lignin peak in the culture broth disappeared.     

Isolation of polyacrylamide-degrading microorganisms from soil

Two polyacrylamide-degrading bacterial strains, No. 2 and No. 11, were isolated from soil, and identified asBacillus sphaericus No. 2 andAcinetobacter sp. No. 11, respectively. Both strains grew on medium containing polyacrylamide as sole carbon and nitrogen sources.B. sphaericus No. 2 andA. sp. No. 11 reduced by 16% and 19% of the initial polyacrylamide concentration, respectively. Optimal pH and temperature in growth ofAcinetobacter sp. No. 11 were 8.0 and 37°C, respectively. After 14-day cultivation ofA. sp. No. 11, the average molecular weight of polyacrylamide has been shifted from 2.3×10⁶ to 0.5×10⁶.     

Exploring the application of biostimulation strategy for bacteria in the bioremediation of industrial effluent

The purpose of this study was to isolate and characterise toxic element-resistant bacteria from acid mine drainage water and to apply them in the bioremediation of industrial effluent, as well as to identify optimal effluent:nutrient concentration for onsite biostimulation strategy. Wastewater samples were collected from acid mine drainage and industry. Inductively coupled plasma optical emission spectroscopy (ICP-OES) was employed for elemental composition analysis. Isolated bacterial strains were characterised using molecular methods. Bioremediation assays were employed to determine the extent of bacterial tolerance and removal of toxic elements using a biostimulation strategy employing minimal salt medium (MSM) at varied concentrations and positive and negative controls of only MSM and industrial effluent, respectively. Two bacterial strains demonstrated resistance to toxic elements, Bacillus sp. MGI101 and Lysinibacillus sp. MGI102 both isolated from the AMD sites. However, no observable growth of toxic metal-resistant bacteria was obtained from the industrial effluents. Bacterial strains MGI101 and MGI102 demonstrated high resistance to target toxic elements during the screening and tolerance assays. Remarkably, Bacillus sp. MGI101 demonstrated greater ability to remove toxic elements including arsenic, chromium, zinc, copper and aluminium in undiluted solutions of the industrial effluent, with its highest removal capacity observed at > 60% for arsenic and aluminium. Both Bacillus sp.MGI101 and Lysinibacillus sp. MGI102 demonstrated varied abilities for the removal of toxic elements from dilution concentration of effluent mixed with MSM. However, the optimal dilution ratio observed in this experiment was 5:15 (effluent:MSM). Overall results demonstrated that isolated bacterial strains have the potential to be employed in bioremediation programmes of acid mine drainages and multi-element contaminated water.