In vitro growth of gelatin suspensions of uredospores of Puccinia graminis f. sp. tritici

1969 ◽  
Vol 47 (8) ◽  
pp. 1291-1293 ◽  
Author(s):  
M. D. Coffey ◽  
A. Bose ◽  
Michael Shaw
Keyword(s):  
Basic Medium ◽  
Australian Isolate ◽  
Brown Pigment ◽  
Puccinia Graminis ◽  
Partial Recovery ◽  
It Diffusion ◽  
The Media ◽  
Agar Surface ◽  
Bacto Agar

A comparison was made of the growth of the Australian isolate of Puccinia graminis Pers. f. sp. tritici Eriks. & E. Henn. race ANZ 126-6,7 on 1.5% Difco Bacto-Agar using water and 15% gelatin as suspension media for seeding uredospores on the media. A range of media was tested, the basic constituents being 3% glucose, Czapek's minerals, peptone, and 1% bovine serum albumin. Spores embedded in gelatin gave much better growth than those applied to the agar surface in water suspension in all media. Sodium citrate (0.2%) inhibited growth; pectin (0.2%) did also, but partial recovery occurred later. Best growth was on the basic medium seeded with uredospores in gelatin. The mycelium was white and fluffy in appearance at first, but collapsed later as a dense brown stroma developed beneath it. Diffusion of brown pigment into the agar medium took place with the gelatin, but not the water series. Sporulation was observed after 60 days in a few colonies and was located in the brown stroma. Spores were non-pigmented, oval in shape, and similar in size to typical uredospores. It is suggested that the physical environment around the spores affects growth.

In vitro growth of wheat and flax rust fungi on complex and chemically defined media

1974 ◽  
Vol 52 (6) ◽  
pp. 1183-1195 ◽  
Author(s):  
A. Bose ◽  
Michael Shaw
Keyword(s):  
Liquid Medium ◽  
Aspartic Acid ◽  
Yeast Extract ◽  
Australian Isolate ◽  
Puccinia Graminis ◽  
Good Growth ◽  
Complex Media ◽  
Mineral Salts ◽  
Melampsora Lini

Growth from uredospores seeded in axenic culture is described for several races of Puccinia graminis Pers. f. sp. tritici (Erikss. and Henn.) and race 3 of Melampsora lini (Ehrenb.) Lév. on complex media containing peptone, yeast extract, and bovine serum albumin (BSA); and for an Australian isolate of Puccinia graminis, race 126-ANZ 6,7, and Melampsora lini, race 3, on chemically defined, liquid media.Of six North American isolates of Puccinia graminis only race 38 formed colonies approaching those of race 126-ANZ 6,7 in final size and general morphology on complex media. 5′AMP had no effect on the growth of 126-ANZ 6,7, but cyclic AMP inhibited growth after uredospore germination. Good growth and sporulation were obtained with 126-ANZ 6,7, but not with the other isolates tested, using a new, chemically defined liquid medium, sterilized by millipore filtration, and containing glucose, Czapek's minerals plus micronutrients, Ca2+, glucose and aspartic acid, glutathione, and cysteine. Uredospores produced in culture reinfected exposed mesophyll tissue, but not intact seedling leaves of wheat.Highly reproducible growth and sporulation of Melampsora lini, race 3, were obtained routinely on a solid medium containing Difco-Bacto agar, sucrose, Knop's minerals, micronutrients, yeast extract, peptone, and BSA. Vegetative cultures, capable of reinfecting the cut ends of surface-sterilized flax cotyledons, could be maintained indefinitely by subdivision before sporulation and transfer to the same medium minus BSA. Evidence is presented that BSA stimulated the development of colonies and the formation of uredospores. The mode of action of BSA is unknown, but it could not be replaced by putrescine.A new chemically defined, liquid medium containing sucrose, Knop's mineral salts, micronutrients, aspartic (or glutamic) acid, and cysteine supported the growth of colonies of Melampsora lini in a highly reproducible manner. The formation of uredospores and teliospores by these colonies was controlled by (a) the level of Ca2+ (as Ca(NO3)2∙4H2O), (b) the concentration of aspartic acid, and (c) the number of colonies per flask. At inoculum levels giving 40 to 60 colonies per flask, in media containing 8.5 mM Ca+ and 45 mM aspartic acid, uredospore formation occurred in 60 to 70% of the colonies. A decrease in the Ca2+ level to 4.25 mM, or a decrease in aspartic acid to 22.5 mM, or adjustment of the inoculum level to give about 10 colonies per flask each resulted in only infrequent sporulation. The uredospores produced in vitro infected intact, 1-week-old flax cotyledons in a normal manner.

scholarly journals Hyaluronan Expression on Vitrified Oocytes Before and After In Vitro Maturation (EKSPRESI HYALURONAN PADA OOSIT YANG DIVITRIFIKASI SEBELUM DAN SESUDAH MATURASI IN VITRO)

2018 ◽  
Vol 19 (1) ◽  
pp. 71
Author(s):  
Zakiyatul Faizah ◽  
R. Haryanto Aswin ◽  
Hamdani Lunardhi ◽  
Widjiati Widjiati
Keyword(s):  
Ethylene Glycol ◽  
In Vitro Maturation ◽  
Cumulus Cells ◽  
Basic Medium ◽  
Oocyte Vitrification ◽  
Temperature Changes ◽  
Before And After ◽  
Rapid Temperature ◽  
The Media

Oocyte vitrification is a major challenge in assisted reproductive technology. Oocyte vitrification with cumulus cells provide benefits in the process of maturation and fertilization. Vitrification leads to rapid temperature changes, therefore the decreasing in temperature could damage the cells even when the morphology was normal. Vitrification of mature oocytes is common because of its low sensitiveness towards low temperatures than immature oocytes. The aim of the research was to compare the effect of vitrification before and after in vitro maturation to the expression of hyaluronan. Maturation was operated in medium TC 50 ?L in CO2 incubators for 24 hours. Vitrification started with washing oocyte in PBS basic medium supplemented with 20% serum for 1-2 minutes, then in equilibration medium PBS + 20% serum + 10% ethylene glycol for 10-14 minutes, then transferred to 20% serum + PBS + 0.5 M sucrose + 15% ethylene glycol + PROH 15% for 25-30 seconds. Thawing was processed by submerging the oocytes in the media: 1). PBS + 20% serum + 0.5 M sucrose (K1); 2) PBS + 20% serum + 0.25 M sucrose (K2); and 3).PBS + 20% serum + 0.1 M sucrose (K3). Immunocytochemical stain was performed to evaluate the hyaluronan expression. Remmele scale index (Immunoreactive score, IRS) was used to read the result. There was no differences of hyaluronan expression in oocyte and cumulus group of K1, K2 and K3 at p< 0.05, statistically. We concluded that there was no difference of hyaluronan expression on oocyte and cumulus between vitrified oocyte of pre and post in vitro maturation which indicated that oocyte could be vitrified in the immature and mature state.

In vitro germination of western larch pollen

2000 ◽  
Vol 30 (2) ◽  
pp. 329-332 ◽  
Author(s):  
Nicole Dumont-BéBoux ◽  
Bradley R Anholt ◽  
Patrick von Aderkas
Keyword(s):  
Pollen Grains ◽  
Major Effect ◽  
Basic Medium ◽  
Tube Length ◽  
In Vitro Germination ◽  
Liquid Media ◽  
Larix Occidentalis ◽  
Western Larch ◽  
The Media

We have been able to successfully germinate western larch (Larix occidentalis Nutt.) pollen in vitro. Pollen was rehydrated at 100% RH for 16 h before being sprinkled on semisolid and liquid media. The basic medium contained Brewbaker and Kwack minerals diluted 1:10 and was supplemented with polyethylene glycol 4000 and three different concentrations of sucrose. The flavonol quercetin was also included in half of the media. More pollen grains survived on liquid media, but semisolid media gave superior germination results. Two to 9% of the grains produced tubes. Quercetin had no major effect on germination, viability, or tube length.

Sporulation and pathogenicity of an Australian isolate of wheat rust grown in vitro

1971 ◽  
Vol 49 (11) ◽  
pp. 1961-1964 ◽  
Author(s):  
A. Bose ◽  
Michael Shaw
Keyword(s):  
Surface Coating ◽  
Germ Tube ◽  
Australian Isolate ◽  
Puccinia Graminis ◽  
Lower Epidermis ◽  
Mineral Salts ◽  
Wheat Leaves ◽  
Wheat Rust ◽  
Serum Albumen

Colonies of Puccinia graminis Pers. f. sp. tritici Erikss. & Henn., race ANZ 126-6,7 were grown from uredospores on Czapek's mineral salts, 3% glucose, 0.1% Evans' peptone, plus defatted bovine serum albumen. Dikaryotic vegetative hyphae apparently developed from centers of germ tube anastomosis, without the formation of typical infection structures. Typical pigmented uredospores and teliospores were formed after 6 to 8 weeks growth. Both spore forms were coated with a layer of material which was visible under the scanning electron microscope and was not observed on uredospores grown on intact wheat leaves. The uredospores were capable of infecting the mesophyll of wheat leaves exposed by stripping back the lower epidermis. The possibility is considered that the surface coating of uredospores grown in vitro is related to their inability to infect intact leaves via the stomata.

scholarly journals Stimulation of development of Cucumis sativus L. plants by low concentration of aluminium in vitro conditions

2013 ◽  
Vol 48 (2) ◽  
pp. 75-82
Author(s):  
Maria Szymańska ◽  
Jolanta Molas
Keyword(s):  
Root System ◽  
Cucumis Sativus ◽  
Leaf Explants ◽  
Small Degree ◽  
Basic Medium ◽  
Cucumis Sativus L ◽  
Leaf Formation ◽  
The Media ◽  
Short Shoots

The culture of cucumber plants <em>Cucumis sativus</em> L. Wisconsin cultivar, from seeds and leaf explants, were carried out on the basic medium ofMurashige and Skoog <em>in vitro</em> conditions. In the culture set from the leaf explants the MS medium was supplemented with IAA (0,5 mg/dm<sup>3</sup>) and BAP (2 mg/dm<sup>3</sup>). Aluminium (as AlCl<sub>3</sub>) was added to the media in concentration Of 1 mg/dm<sup>3</sup>. The media pH was adjusted to 6,2 or 4,2. In seedling culture, aluminium substantially stimulated the growth and development of the root system while a shoot to a small degree only (it advanced leaf formation mainly), causing no morphological and developmental anomalies. In the culture from the leaf explants Al induced the process of rhizogenesis which did not take place on the media without Al. It also stimulated a shoot morphogenesis. After 8 weeks of culture, 32 % leafexplants formed plants with short shoots (2-3,5 cm), long ones (5,5-7 cm) and with long but poorly branched root system„ In the acid conditions (pH 4,2), the effect 0 Al on plant growth was Iower than on the media with pH 6,2. Also a number of regenerated explants with comparable direction of differentiation won a fewer in low pH.

Some influence of the hormonal components in nutrient medium onthe induction of novel structures when obtaining rapeseed haploids through in vitro culture

2018 ◽  
pp. 30-37
Keyword(s):  
Anther Culture ◽  
Hormonal Regulation ◽  
Control Plant ◽  
Pollen Grains ◽  
Control Treatment ◽  
Basic Medium ◽  
Nutrient Media ◽  
Oilseed Crops ◽  
Wide Range

Growth regulators, phytohormones, both natural and artificial, are the main means to control plant ontogenesis. They are involved in regulating the processes of cell differentiation and cell divisions, the formation of tissues and organs, the changes in the rate of growth and development, the duration of the certain stages of ontogenesis. The main classes of phytohormones used in plant biotechnology, in particular, in the induction of haploid structures, are auxins and cytokinins. The mechanism of action of phytohormones on a cell is rather complicated and may have a different character. Understanding the characteristics of the action of phytohormones is complicated by the fact that the system of hormonal regulation of plant life is multicomponent. This is manifested in the fact that the same physiological process is most often influenced not by one, but by several phytohormones, covering a wide range of aspects of cell metabolism. In connection with the foregoing, the purpose of our work was to test a set of nutrient media with different basic composition and different proportions of phytohormones to determine the patterns of their influence on the processes of haploid structure induction in rape anther culture using accessions, developed at the Institute of Oilseed Crops NAAS. The material used was two accessions of winter rapeseed (No. 1 and No. 2) and one sample of spring rapeseed, provided by the Rapeseed Breeding laboratory of the Institute of Oilseed Crops. Incised inflorescences were kept against the background of low temperature of 6–8 ° C for several days, and then, under aseptic conditions, anthers with unripe pollen grains were isolated and planted on nutrient media differing in both basic mineral composition and content of phytohormones. MS (Murashige & Skoog 1962) and B5 (Gamborg et al 1968) media were used as basic media. Phytohormones were added to the basic media in various combinations – BA, 2,4-D, NAA at the concentrations of 0.1-0.6 mg/l. In each treatment up to 300 anthers were cultivated. Differences between treatments were evaluated using standard t-test. Studies have shown that in the anther culture of rapeseed on the tested nutrient media, morphogenic structures of different types (embryoids and callus) were originated. Synthetic auxin 2,4-D, regardless of the composition of the basic medium, caused the formation of structures of both types, though with a low frequency. Phytohormone BA of the cytokinin type had a similar effect. In this case, the frequency of structures was slightly higher, and the developed structures were represented mainly by embryoids. The joint action of cytokinin and auxin was the most favorable for the initiation of morphogenic structures. Such combination of phytohormones caused the formation of these structures with a frequency of 24.5-14.7% in the studied genotypes of winter rape. A similar effect of phytohormones on the induction and development of morphogenic structures was also observed in spring rape. In this case, a single basic MS medium was used. The experiment included treatments where phytohormones were absent (control), as well as various combinations of auxin and cytokinin. In the control treatment, the formation of new structures was not noted. In treatments with phytohormones, in addition to the medium with the combination of auxin and cytokinin, the medium in which only cytokinin was present was also rather effective. The treatment in which the action of auxin 2,4-D was combined with the action of another auxin, NAA, turned out to be practically ineffective. Thus, it was found that for the induction of morphogenic structures from microspores in rape anther culture of the tested genotypes, the combination of cytokinin with auxin, or the use of only single cytokinin BA without other phytohormones, had the most positive effect.

In vitro fertilization in inbred BALB/c mice I: isotonic osmolarity and increased calcium-enhanced sperm penetration through the zona pellucida and male pronuclear formation

Zygote ◽  
2008 ◽  
Vol 16 (3) ◽  
pp. 249-257 ◽  
Author(s):  
Seiji Kito ◽  
Yuki Ohta
Keyword(s):  
Calcium Concentration ◽  
Detrimental Effect ◽  
Choline Chloride ◽  
Positive Control ◽  
Basic Medium ◽  
Vitro Fertilization ◽  
Medium Osmolarity ◽  
Sperm Penetration ◽  
Pronuclear Formation

SummaryTo optimize IVF conditions for BALB/c mice, which are known to have poor in vitro fertilizability, the requirements for sperm–ova interaction were studied by use of modified simplex optimization medium (mKSOM) as a basic medium. Modified human tubal fluid (mHTF) was used for sperm preincubation and acted as a positive control. When the two media were compared, neither capacitation nor fertilization was supported in mKSOM. Increasing the calcium concentration in mKSOM to 5 mM or more during sperm: ova coincubation improved zona penetration but not male pronuclear (MPN) formation to the same level as those cells incubated in mHTF. When medium osmolarity was varied from 230–305 mOsmol by NaCl at 5 mM CaCl2, MPN formation improved at 280 mOsmol or higher osmolarity to the same level as that found when using mHTF. When NaCl equivalent to 25–75 mOsmol was substituted with trehalose, no significant reduction in fertilization was observed. Substitution of NaCl equivalent to 75 mOsmol with other osmotic reagents (sucrose, choline chloride and sorbitol) resulted in similar levels of fertilization as found with mHTF, except for sorbitol, which reduced fertilization significantly caused by its detrimental effect on sperm viability. At isotonic osmolarity (305 mOsmol), maximum fertilization was observed at 5 mM CaCl2; lower or higher concentrations of CaCl2 resulted in reduced fertilization. Calcium and osmolarity, therefore, are important for sperm : ova interaction in BALB/c mice and the increases in calcium to 5 mM and osmolarity to 305 mOsmol are optimal for BALB/c sperm to penetrate through the zona and to form MPN.

scholarly journals Viability of ‘Candidatus Liberibacter asiaticus’ Prolonged by Addition of Citrus Juice to Culture Medium

2014 ◽  
Vol 104 (1) ◽  
pp. 15-26 ◽  
Author(s):  
Jennifer K. Parker ◽  
Sarah R. Wisotsky ◽  
Evan G. Johnson ◽  
Faraj M. Hijaz ◽  
Nabil Killiny ◽  
...  
Keyword(s):  
Grapefruit Juice ◽  
Culture Medium ◽  
Quantitative Polymerase Chain Reaction ◽  
Chemical Characterization ◽  
Candidatus Liberibacter Asiaticus ◽  
Citrus Greening Disease ◽  
Candidatus Liberibacter ◽  
The Media ◽  
Liberibacter Asiaticus

Huanglongbing, or citrus greening disease, is associated with infection by the phloem-limited bacterium ‘Candidatus Liberibacter asiaticus’. Infection with ‘Ca. L. asiaticus’ is incurable; therefore, knowledge regarding ‘Ca. L. asiaticus’ biology and pathogenesis is essential to develop a treatment. However, ‘Ca. L. asiaticus’ cannot currently be successfully cultured, limiting its study. To gain insight into the conditions conducive for growth of ‘Ca. L. asiaticus’ in vitro, ‘Ca. L. asiaticus’ inoculum obtained from seed of fruit from infected pomelo trees (Citrus maxima ‘Mato Buntan’) was added to different media, and cell viability was monitored for up to 2 months using quantitative polymerase chain reaction in conjunction with ethidium monoazide. Media tested included one-third King's B (K), K with 50% juice from the infected fruit, K with 50% commercially available grapefruit juice, and 100% commercially available grapefruit juice. Results show that juice-containing media dramatically prolong viability compared with K in experiments reproduced during 2 years using different juice sources. Furthermore, biofilm formed at the air–liquid interface of juice cultures contained ‘Ca. L. asiaticus’ cells, though next-generation sequencing indicated that other bacterial genera were predominant. Chemical characterization of the media was conducted to discuss possible factors sustaining ‘Ca. L. asiaticus’ viability in vitro, which will contribute to future development of a culture medium for ‘Ca. L. asiaticus’.

Drug activity against Onchocerca gutturosa males in vitro: a model for chemotherapeutic research on onchocerciasis

1987 ◽  
Vol 61 (4) ◽  
pp. 271-281 ◽  
Author(s):  
Simon Townson ◽  
C. Connelly ◽  
A. Dobinson ◽  
R. Muller
Keyword(s):  
Culture System ◽  
Serum Proteins ◽  
Good Condition ◽  
Calf Serum ◽  
New Drugs ◽  
Foetal Calf Serum ◽  
Partial Recovery ◽  
Feeder Cells ◽  
Compound Identification

ABSTRACTAn in vitro system for chemotherapeutic research using adult male Onchocerca gutturosa has been developed as a model for O. volvulus. Using a culture system consisting of medium MEM+10% heat inactivated foetal calf serum (IFCS)+LLCMK2 (monkey kidney) feeder cells in an atmosphere of 5% CO2 in air, we examined the effects of a range of antiparasitic drugs on worm motility. Ivermectin, levamisole, furapyrimidone, Mel W, chloroquine, metrifonate, flubendazole, amoscanate and the Ciba-Geigy compounds CGP 6140, CGP 20′376 and CGI 17658 either immobilized or significantly reduced motility levels at a concentration of 5x10−5M or less within a 7-day period. Worms were affected at very low concentrations by ivermectin (effective conc. to reduce motility levels to 50% of controls, 3.14x10−8M), levamisole (7.95x10−8M), CGP 6140 (8.87x10−9M) and CGP 20′376 (2.78x10−8M). Difficulties were experienced in accurately repeating the immotile endpoint for levamisole due to an inconsistent partial recovery of motility. Over a 7-day period diethylcarbamazine had little effect on motility levels, while suramin caused a slight increase in activity compared to controls at some timepoints. Subsequent experiments demonstrated some differences in drug efficacy depending on the presence or absence of serum and feeder cells in the culture system probably because of drug avidly binding to serum proteins. However, serum and cells were found to be essential ingredients of the culture system to maintain worms in good condition, indicating that new drugs should be evaluated both in the presence and absence of serum and cells. Comparisons were made between the responses of O. gutturosa and Brugia pahangi to certain drugs and these species were found to significantly differ in their sensitivities to ivermectin and a novel compound (Wellcome), indicating that Onchocerca parasites should be used wherever possible for compound identification and development intended for the treatment of onchocerciasis. The in vitro system described here, using male O. gutturosa, provides a basis for further research and a practical alternative to O. volvulus.

Biochemical markers for pregnancy in the spent culture medium of in vitro produced bovine embryos

2021 ◽  
Author(s):  
Gabriela de Oliveira Fernandes ◽  
Marcella Pecora Milazzotto ◽  
Andrei Antonioni Guedes Fidelis ◽  
Taynan Stonoga Kawamoto ◽  
Ligiane de Oliveira Leme ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Biochemical Markers ◽  
Culture Medium ◽  
Culture Media ◽  
Ionization Mass ◽  
Metabolic Profiles ◽  
Bovine Embryos ◽  
The Media

Abstract The present study aimed to identify biomarkers to assess the quality of in vitro produced (IVP) bovine embryos in the culture media. IVP embryos on Day (D) 5 of development were transferred to individual drops, where they were maintained for the last 48 h of culture. Thereafter, the medium was collected and the embryos were transferred to the recipients. After pregnancy diagnosis, the media were grouped into the pregnant and nonpregnant groups. The metabolic profiles of the media were analyzed via electrospray ionization mass spectrometry, and the concentrations of pyruvate, lactate, and glutamate were assessed using fluorimetry. The spectrometric profile revealed that the media from embryos from the pregnant group presented a higher signal intensity compared to that of the nonpregnant group; the ions 156.13 Da [M + H]+, 444.33 Da [M + H]+, and 305.97 Da [M + H]+ were identified as biomarkers. Spent culture medium from expanded blastocysts (Bx) that established pregnancy had a greater concentration of pyruvate (p = 0.0174) and lesser concentration of lactate (p = 0.042) than spent culture medium from Bx that did not establish pregnancy. Moreover, pyruvate in the culture media of Bx can predict pregnancy with 90.9% sensitivity and 75% specificity. In conclusion, we identified markers in the culture media that helped in assessing the most viable IVP embryos with a greater potential to establish pregnancy.

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