Adaptation of stomatal response ofCamellia rusticana to a heavy snowfall environment: Winter drought and net photosynthesis

The adaptation ofCamellia rusticana, an evergreen broad-leaved shrub found in areas of heavy snowfall in Japan, to heavy snowfall environments, and the mechanisms by which it is damaged in winter above the snow, were investigated. The stomatal response and photosynthetic characteristics ofC. rusticana were compared to those ofCamellia japonica found in areas of light snowfall. In field conditions, the mean net photosynthesis ofC. rusticana at photon flux density (PFD) over 200 μmol m⁻²s⁻¹ (Pn(_>200). was 50% larger than that ofC. japonica, but in both light saturated and CO₂ saturated conditions, the O₂ evolution rate (Pc) ofC. rusticana was not different from that ofC. japonica. Mean leaf conductance at PFD over 200 μmol m⁻²s⁻¹ (gl(_>200)) was about 100% larger than that ofC. japonica in the field. The Pn(_>200)) was 50% ratio ofC. rusticana was 37% higher than that ofC. japonica which suggests thatC. rusticana's larger Pn(_>200) can be explained by its larger gl(_>200). WhenC. rusticana trees wintering underneath the snow were projected above it, the leaves of these plants showed serious drought within five days in non-freezing conditions. Their Pc and the maximum stomatal conductance decreased by half and did not recover. The leaves ofC. rusticana showed larger gl(_>200) and a less sensitive stomatal response to the decrease of leaf water potential than that ofC. japonica. The stomata characteristics ofC. rusticana caused larger net photosynthesis than that ofC. japonica during the no snow period, and caused the need for snow cover in winter as protector from winter drought.     

Polyol metabolism in the mycelium and fruit-bodies during development ofFlammulina velutipes

Changes of polyol contents in the mycelium and fruit-bodies ofFlammulina velutipes were measured. The results suggested that arabinitol is accumulated in the fruit-bodies as the end-product after its translocation from the mycelium, while mannitol in the fruit-bodies is converted into fructose by the action of mannitol dehydrogenase (MDH). The development of fruit-bodies was promoted by feeding of mannitol to the mycelial colony. A¹⁴C tracer experiment indicated that half of mannitol translocated from mycelium to fruit-bodies was utilized for fruit-body development. NAD-linked MDH andd-arabinitol dehydroganase (D-ADH) were detected in both mycelium and fruit-bodies. The activities of MDH and ADH in the mycelium reached their maximum levels in the inital stage of fruit-body development and decreased thereafter. In contrast, the activity of MDH in the fruit-bodies showed a peak in the middle stage of development. The activity of ADH in the fruit-bodies was less than half of that of MDH. MDH showed a lower Km value for mannitol (1.3 ×10⁻³M) than for fructose (6.0×10⁻² M). The Km value of ADH for arabinitol was extremely high (1.3×10⁻¹M).     

Effects of concentrated carbon dioxide on the fruiting of several cultivated basidiomycetes (II)

Edible basidiomycetesFlammulia velutipes andPleurotus ostreatus were cultivated in the usual manner on media based on sawdust and rice bran, and the cultures were exposed to slowly flowing CO₂-enriched air (550 (control), 3,000, 6,000, and 9,000µl/l) for seven days at different stages of cultivation. When the cultures were exposed at the primordium stage (less than 10 mm in length), length and yield of fruit-bodies increased and pileus expansion was slightly inhibited inF. velutipes, while inP. ostreatus length increased, yield decreased, and pileus expansion was greatly inhibited. When the cultures with fruit-bodies larger than 10 mm were exposed, length and yield were insensitive and pileus expansion was greatly inhibited inF. velutipes, while inP. ostreatus length was insensitive, but pileus expansion was heavily damaged by trumpet-like deformation and yield decreased. The different action of CO₂ on the two species appeared to be due to the different anatomical structures of their fruit-bodies.     

Enhanced Chlorella vulgaris Buitenzorg growth by photon flux density alteration in serial photobioreactors

Microalgae perform oxygenic photosynthesis and are capable of taking up a large amount of CO₂, using an inducible CO₂ concentrating mechanism (CCM), and fixing CO₂ into higher compounds. These characteristics make the microalgae potentially useful for removal and utilization of CO₂ emitted from industrial plants and, generally, the usage of photosynthetic microorganisms has increased and significantly improved as a solution for CO₂ emissions. In this light and based on previous research using Anabaena cylindrica IAM M1 and Spirulina platensis IAM M 135, enhancement was sought for CO₂ fixation and biomass production by Chlorella vulgaris Buitenzorg by increasing the photon flux density concurrent with increases in culture biomass during the cellular growth phase and was compared to cultures of Chlorella grown at optimal constant illumination, with all cultures grown using Bennick basal medium, 29°C, and a flow of 1.0 atm. 10% CO₂ enriched air delivered to three in serial photobioreactors of 0.200 dm³ capacity each. The results showed that increasing illumination during culture increased biomass production of Chlorella by ∼60% as well as increased CO₂ fixation ability by ∼7.0%. It was also demonstrated that the non-competitive inhibition of [HCO₃ ⁻] as a carbon source significantly affected the cultivation in both the increasing and constant photon flux density regimes.     

Avocado shoot culture, plantlet development and net CO2 assimilation in an ambient and CO2 enhanced environment

Summary The proliferation and survival of avocado nodal cultures of juvenile origin were affected by the form and concentration of nitrogen. Optimum growth was achieved on modified Murashige and Skoog medium containing 67% KNO₃ and 33% NH₄NO₃ with total N of 40 mM supplemented with 100 mg l⁻¹ myo-inositol, 1 mg l⁻¹ thiamine HCl, 30 g l⁻¹ sucrose, and 4.44 μM BA with a 16-h photoperiod (120–150 μmol m⁻² s⁻¹). Proliferating shoots and plantlets were photosynthetically active. Better shoot growth and accumulation of higher biomass occurred in a CO₂-enriched environment than under ambient CO₂ conditions. CO₂ assimilation efficiency, however, was higher under the latter conditions than in a CO₂-enhanced environment, e.g., 31±7 and 17±2 μmol CO₂ m⁻² s⁻¹, respectively. The net CO₂ assimilation rates of in vitro grown plantlets were comparable to those of seedlings ex vitro.     

Changes in low molecular weight carbohydrates composition and chitin throughout the cultivation stages ofPleurotus ostreatus

Changes in contents of soluble low molecular weight carbohydrates and chitin in a sawdust-rice bran medium during mycelial growth ofPleurotus ostreatus in bottle cultivation were examined in relation to fruit-body yield of nine stocks. Glucose, mannitol, inositol, sucrose, and trehalose were detected in cultures after mycelial spreading. No significant correlation was observed between contents of soluble low molecular weight carbohydrate during mycelial growth and the fruit-body yield. Negative correlation was found between trehalose content in post-harvest cultures and the fruit-body yield. Chitin content in cultures decreased in the fruiting stage. Positive correlation was detected between chitin content of fruit-bodies and the decrement of chitin in post-harvest culture caused by fruit-body growth.     

Nitrite utilization by Chaetoceros muelleri under elevated CO2 concentration

Chaetoceros muelleri (Lemn.) was cultured with nitrite (NO₂⁻) or nitrate (NO₃⁻) as the sole nitrogen source and aerated with air or with CO₂-enriched air. Cells of C. muelleri excreted into the medium nitrite produced by reduction of nitrate when grown with 100 μM NaNO₃ as nitrogen source. Accordingly, NO₂⁻ concentration reached 10.4 μM after 95 h at the low CO₂ condition (aerated with air); while the maximum NO₂⁻ concentration was only around 2.0 μM at the high CO₂ condition (aerated with 5% CO₂ in air), furthermore, after 30 h it decreased to no more than 1.0 μM. NO₂⁻ was almost assimilated in 80 h when C. muelleri was cultured at the high CO₂ condition with 100 μM NaNO₂ as sole nitrogen source. At the high CO₂ condition, after 3 h the activity of nitrite reductase was as much as 50% higher than that at the low CO₂ condition. It was indicated that enriched CO₂ concentration could inhibit nitrite excretion and enhance nitrite assimilation by cells. Therefore, aeration with enriched CO₂ might be an effective way to control nitrite content in aquaculture systems.     

Soil O2 and CO2 effects on root respiration of cacti

Through use of a recently developed technique that can measure CO₂ exchange by individual attached roots, the influences of soil O₂ and CO₂ concentrations on root respiration were determined for two species of shallow-rooted cacti that typically occur in porous, well-drained soils. Although soil O₂ concentrations in the rooting zone in the field were indistinguishable from that in the ambient air (21% by volume), the CO₂ concentrations 10 cm below the soil surface averaged 540 μLL⁻¹ for the barrel cactusFerocactus acanthodes under dry conditions and 2400 μLL⁻¹ under wet conditions in a loamy sand. For the widely cultivated platyopuntiaOpuntia ficus-indica in a sandy clay loam, the CO₂ concentration at 10 cm averaged 1080 μLL⁻¹ under dry conditions and 4170 μLL⁻¹ under wet conditions. For both species, the respiration rate in the laboratory was zero at 0% O₂ and increased to its maximum value at 5% O₂ for rain roots (roots induced by watering) and 16% O₂ for established roots. Established roots ofO. ficus-indica were slightly more tolerant of elevated CO₂ than were those ofF. acanthodes, 5000 μLL⁻¹ inhibiting respiration by 35% and 46%, respectively. For both species, root respiration was reduced to zero at 20,000 μLL⁻¹ (2%) CO₂. In contrast to the reversible effects of 0% O₂, inhibition by 2% CO₂ was irreversible and led to the death of cortical cells in established roots in 6 h. Although the restriction of various cacti and other CAM plants to porous soils has generally been attributed to their requirement for high O₂ concentrations, the present results indicate that susceptibility of root respiration to elevated soil CO₂ concentrations may be more important.     

Comparative study of leaf carbon gain in saplings ofThujopsis dolabrata var.hondai andQuercus mongolica var.grosseserrata in a cool-temperate deciduous forest

Seasonal courses of leaf CO₂ gas exchange in a growing season were examined in saplings ofThujopsis dolabrata var.hondai andQuercus mongolica var.grosseserrata in a cool temperate deciduous forest. Between the two tree species there were no large differences in the light compensation point of leaf photosynthesis, except for the season of new leaf expansion. However, light-saturated rates of net photosynthesis were obviously high inT. dolabrata var.hondai. EvergreenT. dolabrata var.hondai saplings had large photosynthetic production in two seasons, before the emergence of new foliage and after foliage fall of the overstory deciduous trees, because of the significantly high solar radiant energy penetrating under the forest canopy during the seasons. Saplings of deciduousQ. mongolica var.grosseserrata were heavily shaded throughout the growing season by foliage of the overstory trees, which resulted in a low daily surplus production. The annual surplus production of leaves in the growing season was estimated to be 2300 mmol CO₂ m⁻² inT. dolabrata var.hondai and −100 mmol CO₂ m⁻², slightly negative, inQ. mongolica var.grosseserrata. These results supported the high survivability ofT. dolabrata var.hondai saplings and the high mortality ofQ. mongolica var.grosseserrata in the deciduous forest.     

CO2 and N-fertilization effects on fine-root length, production, and mortality: a 4-year ponderosa pine study

We conducted a 4-year study of juvenile Pinus ponderosa fine root (≤2 mm) responses to atmospheric CO₂ and N-fertilization. Seedlings were grown in open-top chambers at three CO₂ levels (ambient, ambient+175 μmol/mol, ambient+350 μmol/mol) and three N-fertilization levels (0, 10, 20 g m⁻² year⁻¹). Length and width of individual roots were measured from minirhizotron video images bimonthly over 4 years starting when the seedlings were 1.5 years old. Neither CO₂ nor N-fertilization treatments affected the seasonal patterns of root production or mortality. Yearly values of fine-root length standing crop (m m⁻²), production (m m⁻² year⁻¹), and mortality (m m⁻² year⁻¹) were consistently higher in elevated CO₂ treatments throughout the study, except for mortality in the first year; however, the only statistically significant CO₂ effects were in the fine-root length standing crop (m m⁻²) in the second and third years, and production and mortality (m m⁻² year⁻¹) in the third year. Higher mortality (m m⁻² year⁻¹) in elevated CO₂ was due to greater standing crop rather than shorter life span, as fine roots lived longer in elevated CO₂. No significant N effects were noted for annual cumulative production, cumulative mortality, or mean standing crop. N availability did not significantly affect responses of fine-root standing crop, production, or mortality to elevated CO₂. Multi-year studies at all life stages of trees are important to characterize belowground responses to factors such as atmospheric CO₂ and N-fertilization. This study showed the potential for juvenile ponderosa pine to increase fine-root C pools and C fluxes through root mortality in response to elevated CO₂.     

The optimal growth conditions for the biomass production of Isochrysis galbana and the effects that phosphorus, Zn2+, CO2, and light intensity have on the biochemical composition of Isochrysis galbana and the activity of extracellular CA

The effects of phosphorus, Zn²⁺, CO₂, and light intensity on growth, biochemical composition, and the activity of extracellular carbonic anhydrase (CA) in Isochrysis galbana were investigated. A significant change was observed when the concentration of phosphorus in the medium was increased from 5 μmol/L to 1000 μmol/L affecting I. galbana’s cell density, biochemical composition, and the activity of extracellular CA. Phosphorous concentration of 50 μmol/L to 500 μmol/L was optimal for this microalgae. The Zn²⁺ concentration at 10 μmol/L was essential to maintain optimal growth of the cells, but a higher concentration of Zn²⁺ (≥ 1000 μmol/L) inhibited the growth of I. galbana. High CO₂ concentrations (43.75 mL/L) significantly increased the cell densities compared to low CO₂ concentrations (0.35 mL/L). However, the activity of extracellular CA decreased significantly with an increasing concentration of CO₂. The activity of extracellular CA at a CO₂ concentration of 43.75 mL/L was approximately 1/6 of the activity when the CO₂ concentration was at 0.35 mL/L CO₂. Light intensity from 4.0 mW/cm² to 5.6 mW/cm² was beneficial for the growth, biochemical composition and the activity of extracellular CA. The lower and higher light intensity was restrictive for growth and changed its biochemical composition and the activity of extracellular CA. These results indicate that phosphorus, Zn²⁺, CO₂, and light intensity are important factors that impact growth, biochemical composition and the activity of extracellular CA in I. galbana.     

The effect of headspace renewal in a Temporary Immersion Bioreactor on plantain (Musa AAB) shoot proliferation and quality

The positive effect of ventilation of the culture container on in vitro shoot proliferation and quality was already proven for different species. Hereafter we report on the evolution of the headspace during in vitro culture of plantain in a Temporary Immersion Bioreactor (TIB) on the one hand, and culture on semi-solid medium on the other hand. The CO₂ and C₂H₄ concentration reached a maximum of 12% and 0.45 μl l⁻¹, respectively in the control treatment on semi-solid medium, compared to 5.7% CO₂ and 0.06 μl l⁻¹ C₂H₄ in TIB. The minimal O₂-concentration on semi-solid medium was 15.1%, compared to 19.3% in TIB. The multiplication rate was best in TIB, 6.4 compared to 4.3 in semi-solid conditions, and this was also the case for shoot height (4.3 cm compared to 3.3 cm), and leaf number (2.6 compared to 1.6). Moreover shoots produced on semi-solid medium showed distorted leaves. A typical day-night pattern in CO₂ and O₂ concentration was observed in TIB, as well as on semi-solid medium; this is illustrative for the photosynthetic capacity of the plant material produced in both systems.     

Nitrogen and Carbon Concentrations, and Stable Isotope Ratios in Mediterranean Shrubs Growing in the Proximity of a CO2 spring

Seasonal changes in foliage nitrogen (N) and carbon (C) concentrations and δ¹⁵N and δ¹³C ratios were monitored during a year in Erica arborea, Myrtus communis and Juniperus communis co-occurring at a natural CO₂ spring (elevated [CO₂], about 700 μmol mol⁻¹) and at a nearby control site (ambient [CO₂], 360 μmol mol⁻¹) in a Mediterranean environment. Leaf N concentration was lower in elevated [CO₂] than in ambient [CO₂] for M. communis, higher for J. communis, and dependent on the season for E. arborea. Leaf C concentration was negatively affected by atmospheric CO₂ enrichment, regardless of the species. C/N ratio varied concomitantly to N. Leaves in elevated [CO₂] showed lower δ¹³C, and therefore likely lower water use efficiencies than leaves at the control site, regardless of the species, suggesting substantial photosynthetic acclimation under long-term CO₂-enriched atmosphere. Leaves of E. arborea showed lower values of δ¹⁵N under elevated [CO₂], but this was not the case of M. communis and J. communis foliage. The use of the resources and leaf chemical composition are affected by elevated [CO₂], but such an effect varies during the year, and is species-dependent. The seasonal dependency and species specificity suggest that plants are able to exploit different available water and N resources within Mediterranean sites. This revised version was published online in July 2006 with corrections to the Cover Date.     

Photosynthetic Response of Carrots to Varying Irradiances

Response to irradiance of leaf net photosynthetic rates (P _N) of four carrot cultivars: Cascade, Caro Choice (CC), Oranza, and Red Core Chantenay (RCC) were examined in a controlled environment. Gas exchange measurements were conducted at photosynthetic active radiation (PAR) from 100 to 1 000 μmol m⁻² s⁻¹ at 20 °C and 350 μmol (CO₂) mol⁻¹(air). The values of P _N were fitted to a rectangular hyperbolic nonlinear regression model. P _N for all cultivars increased similarly with increasing PAR but Cascade and Oranza generally had higher P _N than CC. None of the cultivars reached saturation at 1 000 μmol m⁻² s⁻¹. The predicted P _N at saturation (P _Nmax) for Cascade, CC, Oranza, and RCC were 19.78, 16.40, 19.79, and 18.11 μmol (CO₂) m⁻² s⁻¹, respectively. The compensation irradiance (I _c) occurred at 54 μmol m⁻² s⁻¹ for Cascade, 36 μmol m⁻² s⁻¹ for CC, 45 μmol m⁻² s⁻¹ for Oranza, and 25 μmol m⁻² s⁻¹ for RCC. The quantum yield among the cultivars ranged between 0.057–0.033 mol(CO₂) mol⁻¹(PAR) and did not differ. Dark respiration varied from 2.66 μmol m⁻² s⁻¹ for Cascade to 0.85 μmol m⁻² s⁻¹ for RCC. As P _N increased with PAR, intercellular CO₂ decreased in a non-linear manner. Increasing PAR increased stomatal conductance and transpiration rate to a peak between 600 and 800 μmol m⁻² s⁻¹ followed by a steep decline resulting in sharp increases in water use efficiency. This revised version was published online in August 2006 with corrections to the Cover Date.     

Ultrahigh-cell-density culture of a marine green alga Chlorococcum littorale in a flat-plate photobioreactor

To test the feasibility of CO₂ remediation by microalgal photosynthesis, a modified type of flat-plate photobioreactor [Hu et al. (1996) Biotechnol Bioeng 51:51–60] has been designed for cultivation of a high-CO₂-tolerant unicellular green alga Chlorococcum littorale. The modified reactor has a narrow light path in which intensive turbulent flow is provided by streaming compressed air through perforated tubing into the culture suspension. The length of the reactor light path was optimized for the productivity of biomass. The interrelationship between cell density and productivity, as affected by incident light intensity, was quantitatively assessed. Cellular ultrastructural and biochemical changes in response to ultrahigh cell density were investigated. The potential of biomass production under extremely high CO₂ concentrations was also evaluated. By growing C. littorale cells in this reactor, a CO₂ fixation rate of 16.7 g CO₂ l⁻¹ 24 h⁻¹ (or 200.4 g CO₂ m⁻² 24 h⁻¹) could readily be sustained at a light intensity of 2000 μmol m⁻² s⁻¹ at 25 °C, and an ultrahigh cell density of well over 80 g l⁻¹ could be maintained by daily replacing the culture medium. Received: 20 October 1997 / Received revision: 19 December 1997 / Accepted: 24 January 1998     

Field measurement of net photosynthesis of mosses at Langhovde,East Antarctica

Net photosynthesis and dark respiration (CO₂ flux) of Antarctic mosses were measured at Langhovde, East Antarctica, from 9 to 17 January 1988. Moss blocks were taken from communities in the Yukidori Valley (69°14′30″S, 39°46′00″E) at Langhovde. Each block was composed ofCeratodon purpureus andBryum pseudotriquetrum, orB. pseudotriquetrum. The upper part of the block was used to measure net photosynthesis and dark respiration. The net photosynthesis of each sample was measured in the field for one or three days with two infrared CO₂ gas analyzers and an assimilation chamber. The relationships of net photosynthetic rate and dark respiration rate, to the water content of the sample, the intensity of solar radiation and the moss temperature were estimated from the field data. The maximum rate of net photosynthesis was about 4 μmol CO₂ m⁻²s⁻¹ at saturating radiation intensity and at optimum temperature, about 10°C. Environmental features of moss habitats in the Yukidori Valley are discussed in relation to these results.     

Production of inulinase by solid-state fermentation: effect of process parameters on production and preliminary characterization of enzyme preparations

This work was aimed at producing inulinase by solid-state fermentation of sugarcane bagasse, using factorial design to identify the effect of corn steep liquor (CSL) and soybean bran concentration, particle size of bagasse and size of inoculum. Maximum inulinase activity achieved was 250 U per g of dry substrate (gds) at 20% (w/w) of CSL, 5% (w/w) of soybean bran, 1 × 10¹⁰ cells mL⁻¹ and particle size of bagasse in the range 9/32 mesh. The use of soybean bran decreased the time to reach maximum activity from 96 to 24 h and the maximum productivity achieved was 8.87 U gds⁻¹ h⁻¹. The maximum activity was obtained at pH 5.0 and 55.0°C. Within the investigated range, the enzyme extract was more thermostable at 50.0°C, showing a D-value of 123.1 h and deactivation energy of 343.9 kJ gmol⁻¹. The extract showed highest stability from pH 4.5 to 4.8. Apparent K _m and V _max are 7.1 mM and 17.79 M min⁻¹, respectively.     

Characterisation of CO2 and HCO3 − uptake in the cyanobacterium Synechocystis sp. PCC6803

The availability of a complete genome database for the cyanobacterium Synechocystissp. PCC6803 (glucose-tolerant strain) has raised expectations that this organism would become a reference strain for work aimed at understanding the CO₂-concentrating mechanism (CCM) in cyanobacteria. However, the amount of physiological data available has been relatively limited. In this report we provide data on the relative contributions of net HCO₃ ⁻ uptake and CO₂ uptake under steady state photosynthetic conditions. Cells were compared after growth at high CO₂ (2% v/v in air) or limiting CO₂ conditions (20 ppm CO₂). Synechocystishas a very high dependence on net HCO₃ ⁻ uptake at low to medium concentrations of inorganic carbon (Ci). At high Ci concentrations net CO₂ uptake became more important but did not contribute more than 40% to the rate of photosynthetic O₂ evolution. The data also confirm that high Ci cells of Synechocystissp. PCC6803 possess a strong capacity for net HCO₃ ⁻ uptake under steady state photosynthetic conditions. Time course experiments show that induction of maximal Ci uptake capacity on a shift from high CO₂ to low CO₂ conditions was near completion by four hours. By contrast, relaxation of the induced state on return of cells to high CO₂, takes in excess of 230 h. Experiments were conducted to determine if Synechocystissp. PCC6803 is able to exhibit a `fast induction' response under severe Ci limitation and whether glucose was capable of causing a rapid inactivation in Ci uptake capacity. Clear evidence for either response was not found. This revised version was published online in August 2006 with corrections to the Cover Date.     

New insights on toluene biodegradation by Pseudomonas putida F1: influence of pollutant concentration and excreted metabolites

The influence of toluene concentration on the specific growth rate, cellular yield, specific CO₂, and metabolite production by Pseudomonas putida F1 (PpF1) was investigated. Both cellular yield and specific CO₂ production remained constant at 1.0 ± 0.1 g biomass dry weight (DW) g⁻¹ toluene and 1.91 ± 0.31 g CO₂ g⁻¹ biomass, respectively, under the tested range of concentrations (2–250 mg toluene l⁻¹). The specific growth rate increased up to 70 mg toluene l⁻¹. Further increases in toluene concentration inhibited PpF1 growth, although inhibitory concentrations were far from the application range of biological treatment processes. The specific ATP content increased with toluene concentration up to toluene concentrations of 170 mg l⁻¹. 3-Methyl catechol (3-MC) was never detected in the cultivation medium despite being an intermediary in the TOD pathway. This suggested that the transformation from toluene to 3-MC was the limiting step in the biodegradation process. On the other hand, benzyl alcohol (BA) was produced from toluene in a side chain reaction. This is, to the best of our knowledge, the first reported case of methyl monoxygenation of toluene by PpF1 not harboring the pWW0 TOL plasmid. In addition, the influence of 3-MC, BA, and o-cresol on toluene degradation was investigated respirometrically, showing that toluene-associated respiration was not significantly inhibited in the presence of 10–100 mg l⁻¹ of the above-mentioned compounds.     

Long-term cultured callus and the effect factor of high-frequency plantlet regeneration and somatic embryogenesis maintenance in <Emphasis Type="Italic">Zoysia japonica</Emphasis>

In this study, we have demonstrated that Zoysia japonica callus induced from mature seeds can produce high frequencies of plant regeneration and somatic embryogenesis, even following a prolonged period of subculturing. Initial callus cultures were induced from mature seeds of Japanese lawngrass (Z. japonica Steud.) incubated on a medium containing major N₆ medium salts, minor Murashige and Skoog (MS) medium salts, and modified MS medium organic elements supplemented with 3 mg L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.01–0.02 mg L⁻¹ 6-benzyladenine. Compact callus were selected and subcultured monthly on a medium containing 2 mg L⁻¹ 2,4-D, 0.5 mg L⁻¹ kinetin, 500 mg L⁻¹ casein hydrolysate, 500 mg L⁻¹ proline, and 500 mg L⁻¹ myoinositol. Callus maintained in vitro for 18 mo could be induced to regenerate plantlets with a frequency of >90%. By contrast, 36-mo-old callus cultures failed to produce normal shoot regeneration. However, the addition of CuSO₄ to the subculture media maintained >90% regeneration frequencies in such long-term callus cultures. Histological observations revealed that plant regeneration occurred both through somatic embryogenesis and organogenesis pathways. The ability to sustainable regeneration in long-term callus cultures will be valuable to the program of genetic transformation and somaclonal variant selection.