Physiological characteristics of 13 common fungal species in bathrooms

To clarify the factors affecting fungal contamination in bathrooms, the growth of 13 common fungal species (13 isolates) in bathrooms was studied under various environmental conditions. Most of the fungi examined grew on media of 0.01% and 0.05% sodium fatty acid and on media of 0.01% anion surfactant. On media of non-ion surfactant, however, growth varied from species to species. Fungi found commonly in bathrooms can be divided into two groups. The first group, including six species — Cladophialophora boppii, Exophiala spinifera, E. salmonis, Phialophora europaea, Phoma herbarum, and Scolecobasidium constrictum — grew on media of 0.01%, 0.05%, and 0.25% non-ion surfactant, with the latter five species also growing on alkali medium. Most of them did not grow at 33°C, or on media with 10% NaCl, however. Fungi of these six species, identified using DNA and morphological analysis, were common in bathrooms, but not in other indoor environments, for example, in house dust or on windows. The second group contained seven species including Aureobasidium sp., Cladosporium cladosporioides, and Fusarium sp., which were common both in house dust and in bathrooms; they did not grow on media of 0.05% or 0.25% non-ion surfactant, but most grew comparatively fast on normal medium (1/4 PDA), and were able to grow on media with 10% NaCl and also at 33°C. The characteristic fungi found in bathrooms were able to exploit surfactant but were unable to grow well under comparatively dry or high-temperature conditions.     

Oat leaf base: tissue with an efficient regeneration capacity

Summary An efficient short term regeneration system using seedling derived oat (Avena sativa) leaf tissue has been developed. Callus derived from the leaf base showed a higher response of plant regeneration than callus initiated from mesocotyls and more mature parts of the leaves. A correlation between the nuclear DNA content of the donor material, as analysed with flow cytometry, and its ability to form callus was observed. Somatic embryogenesis was histologically recognised from callus derived from tissue close to the apical meristem. Plant regeneration media with various concentrations of auxin were tested. Callus from three different cultivars had a similar regeneration potential with an optimal regeneration frequency of 60%. About 2 months after inoculation regenerated plantlets could be moved to a greenhouse for cultivation.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - DAPI 6-diamidino-2-phenylindole - IAA indole-3-acetic acid - KT kinetin - MS Murashige and Skoog's medium - NAA naphthalene acetic acid     

In vitro culture of lemon juice vesicles

This study explores the possibility of culturing Citrus limon (L.) Burm. f. cv. Eureka (lemon) juice vesicles as isolated intact tissues capable of retaining their unique growth structure in vitro. Isolated juice vesicles either attached to or detached from the endocarp/mesocarp (albedo) tissue of origin were grown on various nutrient media using several physical environments. Various growth responses achieved in vitro from cultured vesicles are described. Intact vesicles attached to endocarp/mesocarp tissue were found to grow up to 6 months in culture as a distinct tissue with a minimum of adverse callus proliferation. Callus formation from some cultured explants occurred on all media and physical environments tested. Callus production eventually obliterated and irreversibly altered the normal development of juice vesicles. Inherent vesicle physiology rather than the tissue culture environment was the major determining factor affecting culture growth. Reducing the concentration of carbohydrates (fructose, glucose or sucrose) added to media from 3 to 0.01% or 0.0% reduced, but still did not totally prevent, callus production. Treatment of vesicles with 1000 mg/l ascorbic acid or citric acid, or 0.1 mg/l indole-3-acetic acid or abscisic acid enhanced the occurrence of normal appearing vesicles.Mention of a trade name or proprietary product or vendor does not constitute approval or guarantee of the product by the U.S. Department of Agriculture, and does not imply its approval to the exclusion of other products that may also be suitable.     

VARIATIONS WITHIN SINGLE CELL CLONES OF TOBACCO TISSUE CULTURES

Comparative studies were made utilizing two series of secondary clones (single cell clones derived from single cell clones H 196 and H 241) of hybrid tobacco (Nicotiana tabacum ♂ × Nicotiana glutinosa ♀ ) tissue grown in vitro. Secondary clones derived from a single parent varied in color, consistency, the ability to grow, and rate of growth with various carbohydrates and growth-promoting substances. The growth of the secondary clones generally resembled that of the parent clone from which derived. Many of the 23 secondary clones of H 196 grew satisfactorily on media supplemented with sucrose, dextrose, levulose, or maltose; lactose, galactose, and xylose were unsatisfactory supplements. Similarly, the series of 30 secondary clones isolated from H 241 grew well on some media but poorly on others. Growth generally decreased when α-napthaleneacetic acid or 2,4-dichlorophenoxyacetic acid was omitted from the basal coconut milk medium. Growth decreased considerably when coconut milk was omitted from the basal medium. The optimum sugar concentration was 1/2 to 1 per cent.     

Shoot Differentiation in Callus Cultures of Datura innoxia

Datura innoxia Mill. callus cultures formed shoots in 2–4 weeks on media containing; a) gibberellic acid, b) indoleacetic acid, c) low concentrations of naphthylacetic acid, d) low concentrations of 2,4-dichlorophenoxyacetic acid, e) benzylaminopurine, f) no growth substance. Benzylaminopurine promoted shoot differentiation. Gibberellic acid inhibited shoot formation weakly, but inhibited proper leaf blade formation. Root differentiation was rare. The callus cultures of Datura innoxia grew rapidly (100-fold in 4 weeks) on a slightly modified Murashige and Skoog medium (0.5 mg/l thiamin · HCl, pH 5.5, no glycine) in light at 30°C. Callus grew well on any single one of the growth substances NAA (10^?5M), 2,4-D (10^?6M) or BAP (3 × 10^?6M). Growth was less and more erratic on GA or IAA. The callus cultures did not grow significantly better when BAP was combined with one of the auxins or with GA.     

Role of origin and endophyte infection in browning of bud-derived tissue cultures of Scots pine (Pinus sylvestris L.)

Callus tissues originating from buds of mature Scots pine (Pinus sylvestris L.) trees exhibit the typical problem of browning, which leads to degeneration and death of the tissues. The effects of medium, origin (tree and location) and endophyte infection were studied on the browning and growth of bud-derived tissue cultures. The calli growing on medium with higher kinetin content and source of organic nitrogen, and originating from the southern location grew better and exhibited less browning. Endophytic microbial cells were detected in the brown callus tissues by transmission electron microscopy. The natural endophyte infection frequency of Scots pine buds was studied and found dependent on the tree, but not on the location. A well-growing, green callus line was artificially infected by an endophytic strain of Methylobacterium extorquens, and browning was not observed on solid media compared to the uninfected control clones of the same callus. However, suspension cultures started from the infected callus died faster than cultures started from the uninfected callus. The endophyte species composition and plant genotype together with tissue culture conditions are the key factors for gaining plant tissue cultures with high regeneration capacity.     

Growth of Avocado Fruit Callus and Its Relation to Exogenous and Endogenous Cytokinins

Callus derived from avocado cotyledons and mesocarp was grown in tissue culture. Cytokinins in nutrient media were vital for the growth of mesocarp callus but not for the growth of cotyledonous callus. It was found that cotyledonous callus synthesized Cytokinins.     

Callus formation in Ecklonia cava Kjellman (Laminariales,Phaeophyta)

Explants from stipes and meristems of Ecklonia cava were incubated on six media under several culture conditions. Both stipe and meristem explants developed calluses three to six weeks after inoculation onto all media except AS PC-1. Calluses developed on stipe explants but did not develop on meristem explants at a temperature of 23 °C. Temperatures from 8 to 13 °C were favorable for callus development. Callus development on meristem explants required light but callus development on stipe explants did not.     

Effects of Pseudomonas syringae pv. morsprunorum Strain C28, Outer Membrane and LPS Preparations on Cherry Callus

Callus cultures were established from cherry (Prunus avium) cvs. Napoleon and Colt, respectively susceptible and resistant to race‐1 strains of Pseudomonas syringae pv. morsprunorum, by growth on Schenk–Hildebrandt medium. On Napoleon callus, necrosis began earlier and proceeded more rapidly when inoculated with the virulent race‐1 cherry isolate strain C28, than with mutants of diminished virulence derived from it, or with the virulent plum isolates D10 and D17. Colt tissue displayed poorer viability and showed susceptibility to strain C28 and the plum isolates. Callus from both sources was somewhat susceptible to the saprophytes P. aeruginosa NCIMB 8295 and P. fluorescens NCIMB 3756. Strain C28 grew on suspended Napoleon callus cells over a period of 3–4 days, causing leakage of UV‐absorbing compounds and K⁺, with a concomitant rise in extracellular pH. P. fluorescens NCIMB 3756 showed no growth on suspended callus for 6 days. EDTA‐extracted outer membrane (OM) from strain C28 caused leakage of UV‐absorbing material and K⁺, which was later reabsorbed, with little change in pH. The presence of OM suppressed the growth of a subsequent inoculum of strain C28, possibly due to complexation of the available Ca²⁺ and/or Mg²⁺ in the surrounding medium, by the component lipopolysaccharide (LPS). OM from the rough avirulent mutant strain C28‐2 induced leakage of K⁺, but not of UV‐absorbing material, and did not prevent the growth of subsequently inoculated strain C28. Smooth LPS from strain C28 did not cause leakage of K⁺ or of UV‐absorbing material and did not prevent growth of C28. The relevance of these findings is discussed in relation to disease.     

The apparent loss of tissue culture competence during leaf differentiation in yams (Dioscorea bulbifera L.)

Explants taken from the leaves of yams (Dioscorea bulbifera L.) at different stages of development were cultured in vitro on a checkerboard using various combinations and/or concentrations of auxin (2,4-d) and cytokinin (6-BAP). An addition of cytokinin to the culture media was not essential for callus induction from explants derived from young leaves in the very early stages of expansion. When the leaves expanded further they required cytokinin and the requirement increased considerably during expansion. Explants taken from fully expanded leaves were no longer able to proliferate, even when extremely high concentrations of cytokinins were applied. Callus grown from highly immature leaves was able to continue proliferating in the absence of cytokinin when subcultured. Callus, that initially required cytokinin in the medium, proliferated in the absence of exogenous cytokinin when subcultured.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - 6-BAP 6-benzylamino purine - 1-NAA 1-naphthalenacetic acid     

CULTURE OF MATURE ECOTYLEDONOUS EMBRYOS OF PHASEOLUS VULGARIS AND THE NUTRITIONAL ROLE OF COTYLEDONS

Embryos of Phaseolus vulgaris L. were excised from seeds and cultured with cotyledons removed to determine the actions of various cultural conditions upon embryo development. Four media were tested, but ecotyledonized embryos did not grow as rapidly on any of them as did embryos with intact cotyledons on agar-water media. Comparisons of growth of ecotyledonized embryos with embryos bearing fractions of cotyledons indicated ecotyledonized embryos cultured on nutrient media grew about as well as embryos bearing cotyledons from which 97% of the volume had been removed surgically. The final weight of ecotyledonized embryos was greater when detached cotyledons were placed near them and was even greater when extracts of detached and incubated cotyledons were employed in the nutrient medium. Benzyladenine, kinetin, gibberellic acid, indole-acetic acid, presence of sucrose, and light or dark culture failed to enhance the ability of incubated cotyledons to stimulate growth of embryos.     

A Comparison of the Effects of Temperature on the Growth of Fusarium oxysporum f. sp. narcissi in Solid and Liquid Media

The effect of different temperatures on mycelial growth of Fusarium oxysporum f. sp. narcissi in liquid and on solid medium was compared. Differences between optimum and maximum temperatures using these substrates were considerable and, could lead to different interpretations of the parasitic growth of the pathogen in vascular tissues, for example, in liquid culture Fusarium grew at 45°C whereas it did not on solid media. Initially F. oxysporum f. sp. narcissi may, be encouraged to grow faster in warm air drying systems than those using lower temperatures. The rates of diffusion between 17°C and 35°C are not markedly different. The speed with which moisture is removed from the internal tissues of the remaining roots is governed more by air flow over the bulbs than by temperature.     

Nitrogen fixation and nitrate utilization by marine and freshwater Beggiatoa

Four newly isolated marine strains of Beggiatoa and five freshwater strains were tested for nitrogen fixation in slush agar medium. All strains reduced acetylene when grown microaerobically in media containing a reduced sulfur source and lacking added combined nitrogen. The addition of 2 mmol N, as nitrate or ammonium salts, completely inhibited this reduction. Although not optimized for temperature or cell density, acetylene reduction rates ranged from 3.2 to 12 nmol·mg prot^-1 min^-1. Two freshwater strains did not grow well or reduce acetylene in medium lacking combined nitrogen if sulfide was replaced by thiosulfate. Two other strains grew well in liquid media lacking both combined nitrogen and reduced sulfur compounds but only under lowered concentrations of air. All freshwater strains grew well in medium containing nitrate as the combined nitrogen source. Since they did not reduce acetylene under these conditions, we infer that they can assimilate nitrate.     

Wheat Callus Culture: the Initiation, Growth and Organogenesis of Callus Derived from Various Explant Sources

Callus was obtained from mature excised embryos of wheat, fromnodal and internodal stem segments and from rachis segmentsusing the medium of Murashige and Skoog(1962)(M medium), containing1-0mg l^–1 2,4-D, and from immature embryos using the mediumof Green and Phillips (1975) containing 2 mg l^–1 2,4-D.Callus yield from mature embryos depended upon the cultivarused. No callus could be obtained from leaf segments. Callusderived from mature embryos and nodal stem segments was successfullymaintained by serial sub-culture on the M medium containing2,4-D for up to 3 years although its growth rate declined toa lower level as culture proceeded. Such cultures consistently produced roots when transferred toa medium containing a low level of 2,4-D or no 2,4-D. The presenceof the auxin was essential for continued proliferation of thecallus tissue. Shoot initiation was infrequent, did not occurafter the first few sub-cultures and could not be enhanced byvarious auxin and cytokinin additions to the medium. Callusderived from immature embryos did not have an enhanced potentialfor shoot initiation. Triticum aestivum, wheat, callus culture, organogenesis     

Growth of cells from esophageal squamous cell carcinoma biopsies in a reconstituted basement membrane

Cells from esophageal carcinoma biopsies were cultured on or inside a three-dimensional basement membrane matrix (matrigel). Their growth was compared to cells derived from control esophageal biopsies. Cells from both normal and neoplastic tissue attached poorly to tissue culture plastic. Matrigel coating improved adhesion and growth. When cells were grown inside a matrigel matrix, a striking difference was noticed between carcinoma cells and controls. Carcinoma cells grew invasively in the three-dimensional substrate and digested the matrix after a few weeks; control cells did not grow and only a few necrotic cells were visible with time. Matrigel provided a better growth substrate than plastic for esophageal derived cells and discriminated between carcinoma-derived and control cells when used as a three-dimensional growth substrate. Our studies suggest a possible use of matrigel for the selective growth of tumor cells derived directly from tissue biopsies.     

Ecological relationships between xerophilic fungi and house-dust mites (Acarida: Pyroglyphidae)

Summary At. 75 and 80% relative humidity (RH), on a wheat germ flake medium, Aspergillus penicilloides grew abundantly and suppressed the population growth of Dermatophagoides pteronyssiunus. At 71% RH, A. penicilloides grew moderately and was only antagonistic to D. pteronyssinus when the fungus was previously incubated on the medium.On a human dander medium and on mattress dust, A. penicilloides grew moderately at 71% and 75% RH and stimulated the development of D. pteronyssinus populations. Also a moderate growth of Eurotium repens on human dander positively influenced D. pteronyssinus. Wallemia sebi and Penicillium brevicompactum grew slightly or did not grow at all at 75% RH. No effect was observed on D. pteronyssinus.It appears that xerophilic fungi may stimulate, and occasionally may reduce, the growth of house-dust mite populations in the natural environment.     

Tissue Culture of Maize I. Callus Induction and Growth

Callus induction and subculture was successful with mature embryos and stem sections of seedlings of Zea mays L. on Linsmaier and Skoog's medium modified to contain 4 mg/I of 2,4-D and 1 g/I of casamino acids. — 2,4-D was superior to NAA and IAA for both callus induction and growth. Callus subcultured on NAA formed abundant roots on agar-solidified media and numerous root-like primordia in liquid cultures. — Kinetin had no effect on callus induction in the presence of 2,4-D and neither kinetin nor gibberellic acid stimulated callus growth during subculture. — Callus grew equally well on the medium of Linsmaier and Skoog, that of Schenk and Hildebrandt, and the B-5 medium of Gamborg and Eveleigh containing 2% sucrose, 4 mg/I of 2,4-D and 1 g/I of casamino acids. — The callus grew more rapidly at 25°C than at 30°C or 35°C. Little difference was noted at any temperature in callus growth in alternating light (16 h) and dark (8 h) or continuous dark. — Sucrose was superior to glucose and maltose in both liquid and agar-solidified cultures. Lactose and galactose failed to support callus growth.     

β-Galactosidase activity in cultured cotton cells (Gossypium Hirsutum L.): A comparison between cells growing on sucrose and lactose

Summary Cotton callus and suspension cultures developed from a cotton variety susceptible toXanthomonas malvacearum (E. F. Sm.) Dow, were grown on chemically defined media that contained one of the carbohydrate sources: 3% sucrose, 3% lactose, 3% maltose, 3% fructose, and 3% glucose. All cells were maintained on a medium with sucrose as the carbohydrate and subsequently transferred to media containing the above carbohydrates. Sucrose was the best carbon source for a high growth rate; however, cells growing on glucose, which was almost as good as sucrose, and cells growing on lactose did not turn brown when they reached the stationary phase of growth. A crude extract from callus tissue growing on lactose has a fivefold increase in β-galactosidase [EC 3.21.23] activity as compared with the extract from callus tissue growing on sucrose. When callus tissue growing on lactose was transferred tomedium containing sucrose, β-galactosidase activity decreased to the level as measured in cells maintained on sucrose. Callus cells growing on a lactose medium showed staining when treated with 5-bromo-4-chloro-3-indolyl β-D-galactopyranoside in which very heavy granular stains appeared. Cells growing on sucrose did not show the histochemical staining. These observations suggest that β-galactosidase is induced in cotton callus tissue that has been transferred from a medium containing sucrose to a medium containing lactose. This is journal article J-3704 of the Oklahoma Agricultural Experiment Station. The research was supported in part by a Presidential Challenge Grant from Oklahoma State University and the Oklahoma Agricultural Experiment Station.     

Selection and characterization of ethionine-resistant alfalfa (Medicago sativa L.) cell lines

Summary Diploid alfalfa (HG2), capable of plant regeneration from tissue culture, was used to select variant cell lines resistant to growth inhibition due to ethionine (an analog of methionine). Approximately 10⁷ suspension-cultured cells were mutagenized with methane sulfonic acid ethylester and then plated in solid media containing ethionine. Callus colonies formed on media with 0.02 mM ethionine. Of the 124 cell lines recovered, 91 regenerated plants. After six months growth on media without ethionine, 15 of 110 cell lines of callus grew significantly better than HG2 on 1 mM ethionine. Several ethionine-resistant callus cultures were also resistant to growth inhibition due to the addition of lysine + threonine to the media. High concentrations, relative to unselected HG2 callus, of methionine, cysteine, cystathionine, and glutathione were found in some, but not all, ethionine-resistant callus cultures. Cell line R32, which had a ca. tenfold increase in soluble methionine, had a 43% increase in total free amino acids and a 40% increase in amino acids in protein as compared to unselected HG2 callus. Relative amounts of each amino acid in protein were the same in both.Abbreviation LT lysine + threonine in equimolar concentration     

Reduction of ferric leghemoglobin in soybean root nodules

Callus tissue cultures were developed from apical meristem regions of tumor-like ineffective root nodules of alfalfa. Callus growth was a function of tissue source and hormone composition and concentration. Callus derived from ineffective nodules also were shown not to contain Rhizobium meliloti.

Glutamate dehydrogenase, glutamine synthetase, glutamate synthase, glutamate oxaloacetate transaminase and phosphoenolpyruvate carboxylase activities were present in callus cultures and in the respective nodule source used for callus induction. The mean specific activity of all enzymes evaluated was higher in callus cultures than in ineffective nodules. Quantitative but not qualitative differences in enzyme activities were evident between ineffective nodules and callus derived from these nodules. Tissue cultures derived from ineffective nodules may provide a model system to evaluate host plant-Rhizobium interactions.