<Emphasis Type="Italic">Trichoderma harzianum</Emphasis>: a biocontrol agent against <Emphasis Type="Italic">Bipolaris oryzae</Emphasis>

Rice brown spot, caused by Bipolaris oryzae, can be a serious disease causing a considerable yield loss. Trichoderma harzianum is an effective biocontrol agent for a number of plant fungal diseases. Thus, this research was carried out to investigate the mechanisms of action by which T. harzianum antagonizes Bipolaris oryzae in vitro, and the efficacy of spray application of a spore suspension of T. harzianum for control of rice brown spot disease under field conditions. In vitro, the antagonistic behavior of T. harzianum resulted in the overgrowth of B. oryzae by T. harzianum, while the␣antifungal metabolites of T.␣harzianum completely prevented the linear growth of B. oryzae. Light and scanning electron microscope (SEM) observations showed no evidence that mycoparasitism contributed to the aggressive nature of the tested isolate of T. harzianum against B. oryzae. Under field conditions, spraying of a spore suspension of T. harzianum at 10⁸ spore ml⁻¹ significantly reduced the disease severity (DS) and disease incidence (DI) on the plant leaves, and also significantly increased the grain yield, total grain carbohydrate, and protein, and led to a significant increase in the total photosynthetic pigments (chlorophyll a and b and carotenoids) in rice leaves.     

Attenuation of cadmium toxicity in mycorrhizal celery (<Emphasis Type="Italic">Apium graveolens</Emphasis> L.)

A pot-culture experiment was carried out to investigate the effect of arbuscular mycorrhizal (AM) fungus (Glomus macrocarpum Tul. and Tul.) on plant growth and Cd²⁺uptake by Apium graveolens L. in soil with different levels of Cd²⁺. Mycorrhizal (M) and non-mycorrhizal (NM) plants were grown in soil with 0, 5, 10, 40 and 80 Cd²⁺ mg kg⁻¹soil. The infectivity of the fungus was not affected by the presence of Cd²⁺ in the soil. M plants showed better growth and less Cd²⁺ toxicity symptoms. Cd²⁺ root : shoot ratio was higher in M plants than in NM plants. These differences were more evident at highest Cd²⁺ level (80 mg kg⁻¹ soil). Chlorophyll a and chlorophyll b concentrations were significantly higher in AM-inoculated celery leaves. The dilution effect due to increased biomass, immobilization of Cd²⁺ in root and enhanced P-uptake in M plants may be related to attenuation of Cd²⁺toxicity in celery.     

Insecticide-tolerant and plant growth promoting <Emphasis Type="Italic">Bradyrhizobium</Emphasis> sp. (<Emphasis Type="Italic">vigna</Emphasis>) improves the growth and yield of greengram [<Emphasis Type="Italic">Vigna radiata</Emphasis> (L.) Wilczek] in insecticide-stressed soils

This study was designed to identify rhizobial strains specific to greengram expressing higher tolerance against insecticides, fipronil and pyriproxyfen, and synthesizing plant growth regulators even amid insecticide-stress. Of the 50 bradyrhizobial isolates, the Bradyrhizobium sp. strain MRM6 showed tolerance up to 1,600 μg mL⁻¹ against each of fipronil and pyriproxyfen. The tolerant Bradyrhizobium sp. (vigna) produced plant growth promoting substances in substantial amounts, both in the presence and absence of insecticides. The strain MRM6 was further used to investigate its impact on greengram grown in soils treated with 200 (the recommended dose), 400 and 600 μg kg⁻¹ soil of fipronil and 1,300 (the recommended dose), 2,600 and 3,900 μg kg⁻¹ soil of pyriproxyfen. Fipronil at 600 μg kg⁻¹ soils and pyriproxyfen at 3,900 μg kg⁻¹ soils had greatest toxic effects and decreased plant biomass, symbiotic efficiency, nutrient uptake and seed yield of greengram plants. The Bradyrhizobium sp. (vigna) inoculant when used with fipronil and pyriproxyfen significantly increased the measured parameters compared to the plants grown in soils treated solely with the same concentration of each insecticide. This study inferred that the Bradyrhizobium sp. (vigna) strain MRM6 may be exploited as bio-inoculant to increase the productivity of greengram exposed to insecticide-stressed soils.     

<Emphasis Type="Italic">Trichoderma harzianum</Emphasis>: occurrence in the air and clinical significance

Trichoderma harzianum, a filamentous fungus, is being widely used as a potential biopesticide. The potential of this fungus in causing skin sensitization, however, was poorly investigated as yet. The objective of this study was to monitor the occurrence of T. harzianum in the air and to explore its skin sensitizing potential. Seasonal periodicity of T. harzianum was studied for the years 2002–2004 by an Andersen air sampler. The skin sensitizing potential of T. harzianum extract was studied in 389 patients with suspected respiratory allergy by skin prick test (SPT) and specific IgE level was determined by ELISA. SDS–PAGE and immunoblotting were also performed. T. harzianum colony count varied from 3.69 to 134.88 CFU m⁻³ with the peak achieved in February. Relative humidity was found to be a significant (P < 0.05) factor predicting the occurrence of T. harzianum in the air. Positive skin reaction (wheal diameter ≥ 3 mm) was observed in 105 patients (26.99%). T. harzianum crude extract was resolved in 18 protein bands (12–72 kDa) on SDS–PAGE (12% gel) including two IgE-binding protein bands (21 and 32 kDa). T. harzianum can be considered an important inhalant allergen.     

<Emphasis Type="Italic">In vitro</Emphasis> flowering of <Emphasis Type="Italic">Perilla frutescens</Emphasis>

This study investigated the factors affecting in vitro flowering of Perilla frutescens. The shoots regenerated from cotyledonary and hypocotyl explants cultured on Murashige and Skoog (MS) medium supplemented with benzyladenine (BA) and indole-3-acetic acid, each at 0.5 mg l⁻¹, were excised and transferred to MS medium containing 30 g l⁻¹ of sucrose, 8.25 g l⁻¹ of ammonium nitrate, and 1.0 mg l⁻¹ of BA. After 40 d of culture, 86.2% of shoots flowered and most of which self-fertilized in vitro and produced mature fruits with viable seeds. These seeds were germinated and plants were grown to maturity and flowered in soil under greenhouse conditions. The in vitro flowering system reported in this study may facilitate rapid breeding of P. frutescens and offers a model system for studying the physiological mechanism of flowering.     

<Emphasis Type="Italic">In vitro</Emphasis> multiplication of heavy metals hyperaccumulator <Emphasis Type="Italic">Thlaspi caerulescens</Emphasis>

A micropropagation protocol through multiple shoot formation was developed for Thlaspi caerulescens L., one of the most important heavy metals hyperaccumulator plants. In vitro seed-derived young seedlings were used for the initiation of multiple shoots on Murashige and Skoog (MS) medium with combinations of benzylaminopurine (BA; 0.5–1.0 mg dm⁻³), naphthaleneacetic acid (NAA; 0–0.2 mg dm⁻³), gibberellic acid (GA₃; 0–1.0 mg dm⁻³) and riboflavin (0–3.0 mg dm⁻³). The maximum number of shoots was developed on medium containing 1.0 mg dm⁻³ BA and 0.2 mg dm⁻³ NAA. GA₃ (0.5 mg dm⁻³) in combination with BA significantly increased shoot length. In view of shoot numbers, shoot length and further rooting rate, the best combination was 1.0 mg dm⁻³ BA + 0.5 mg dm⁻³ GA₃ + 1.0 mg dm⁻³ riboflavin. Well-developed shoots (35–50 mm) were successfully rooted at approximately 95 % on MS medium containing 20 g dm⁻³ sucrose, 8 g dm⁻³ agar and 1.0 mg dm⁻³ indolebutyric acid. Almost all in vitro plantlets survived when transferred to pots.     

Intercropping <Emphasis Type="Italic">Caragana arborescens</Emphasis> with <Emphasis Type="Italic">Salix miyabeana</Emphasis> to Satisfy Nitrogen Demand and Maximize Growth

Willow shows great promise as a biomass crop and is now used worldwide. However, willow is a nutrient and water demanding plant that often requires the use of nitrogen (N) fertilizer to maximize growth on poor soils. The intercropping of Salix miyabeana with the atmospheric N₂-fixing Caragana arborescens on poor soils of the Canadian Prairies could provide a portion of the N demand of the willow. The main objectives were to: (1) determine the yield potential, N nutrition and water use efficiency (WUE) of willow and Caragana grown in pure and mixed plantations across a range of soil productivity and (2) assess the extent of atmospheric N₂-fixation by the Caragana within the first rotation in central Saskatchewan. We found large differences in willow yields, foliar N and WUE across the sites. The willow yields (1.24 to 15.6 t dry matter ha⁻¹ over 4 years) were low compared to northeastern North American values and reflect the short and dry summers of the region. The yields were positively correlated to foliar N (ranging between 14.3 and 32.4 mg g⁻¹), whereas higher WUE (expressed as δ¹³C) were not positively correlated to water availability but to higher yields. Caragana N₂-fixation (measured using ¹⁵N isotope dilution) was not active at the most productive site but up to 60% of the foliar N was of atmospheric origin at the two other sites. Willow growth increased with Caragana proportions at the least productive site, which is typical of the benefits of N₂-fixing plants on the growth of other plants on poor soils. At the most productive site, Caragana decreased the growth of willow early on due to competition for resources, but willow eventually shaded Caragana to a point of significant canopy decline and dieback. It is therefore more appropriate to intercrop the two species on less productive soils as Caragana is more likely to add N to the system via N₂-fixation and is less likely to be shaded out by willow.     

Improved Tolerance of <Emphasis Type="Italic">Acacia nilotica</Emphasis> to Salt Stress by Arbuscular Mycorrhiza, <Emphasis Type="Italic">Glomus fasciculatum</Emphasis> may be Partly Related to Elevated K/Na Ratios in Root and Shoot Tissues

A pot experiment was conducted to examine the effect of arbuscular mycorrhizal fungus, Glomus fasciculatum, and salinity on the growth of Acacia nilotica. Plants were grown in soil under different salinity levels (1.2, 4.0, 6.5, and 9.5 dS m⁻¹). In saline soil, mycorrhizal colonization was higher at 1.2, 4.0, and 6.5 dS m⁻¹ salinity levels in AM-inoculated plants, which decreased as salinity levels further increased (9.5 dS m⁻¹). Mycorrhizal plants maintained greater root and shoot biomass at all salinity levels compared to nonmycorrhizal plants. AM-inoculated plants had higher P, Zn, and Cu concentrations than uninoculated plants. In mycorrhizal plants, nutrient concentrations decreased with the increasing levels of salinity, but were higher than those of the nonmycorrhizal plants. Mycorrhizal plants had greater Na concentration at low salinity levels (1.2, 4.0 dS m⁻¹), which lowered as salinity levels increased (6.5, 9.5 dS m⁻¹), whereas Na concentration increased in control plants. Mycorrhizal plants accumulated a higher concentration of K at all salinity levels. Unlike Na, the uptake of K increased in shoot tissues of mycorrhizal plants with the increasing levels of salinity. Our results indicate that mycorrhizal fungus alleviates deleterious effects of saline soils on plant growth that could be primarily related to improved P nutrition. The improved K/Na ratios in root and shoot tissues of mycorrhizal plants may help in protecting disruption of K-mediated enzymatic processes under salt stress conditions.     

Removal of <Emphasis Type="Italic">p</Emphasis>-chlorophenol by the marine microalga <Emphasis Type="Italic">Tetraselmis marina</Emphasis>

The ability of Tetraselmis marina, a green coastal microalga, to remove chlorophenols under photoautotrophic conditions was investigated. T.marina was able to grow in the presence of 20 mg L⁻¹ of the phenolic compounds tested. The EC₅₀ (growth rate) value of p-chlorophenol (p-CP) to T.marina was found to be 25.5 mg L⁻¹. The microalga was able to remove chlorophenols, showing higher efficiency for p-CP. The effect of photoregime and NaHCO₃ concentration on p-CP removal was investigated. Under continuous illumination with 1 g L⁻¹ NaHCO₃ initial concentration T.marina removed 65% of 20 mg L⁻¹ in a 10-day cultivation period.     

Toxic metals accumulation in <Emphasis Type="Italic">Trichoderma asperellum</Emphasis> and <Emphasis Type="Italic">T. harzianum</Emphasis>

Heavy metal contamination represents an important environmental issue due to the toxic effects of metals on different organisms. Filamentous fungi play an important impact in the bioremediation of heavy metal-contaminated wastewater and soil. The purpose of this investigation was to observe fungal uptake behavior toward heavy metal. For this aim Trichoderma asperellum TS141 and T. harzianum TS103 at growth period were screened for their tolerance and uptake capability of cadmium (Cd), lead (Pb) and nickel (Ni) at different concentrations (0, 25, 50, 100, and 200 mg/L) in PDB media (potato dextrose broth as a complex medium). Results showed that both fungi were able to survive at the maximum concentration of 200 mg/L of the heavy metals, and remove them. T. asperellum had a better uptake capacity for Cd compared to Pb and Ni in the highest metal concentration in media. Maximum removal efficiency of Pb (68.4%) at 100 mg/L and Ni (78%) at 200 mg/L was performed by T. asperellum. For Cd, the highest removal efficiency (82.1%) was recorded by T. harzianum at 200 mg/L Cd in aqueous solution. The uptake of Cd was highly dependent on pH of solution than Pb and Ni so that the optimal pH of Cd uptake was 9 for T. asperellum and 4 for T. harzianum. Also, optimal temperature was 35°C for Cd and Pb uptake in both fungi, whereas for Ni uptake was 30 and 35°C in T. harzianum and T. asperellum, respectively. We propose that T. asperellum TS141 and T. harzianum TS103 can be used as a bioremediation agent for metal remediation from wastewater and heavy metal-contaminated soils.     

Carbon nutrition of mature green orchid <Emphasis Type="Italic">Serapias strictiflora</Emphasis> and its mycorrhizal fungus <Emphasis Type="Italic">Epulorhiza</Emphasis> sp.

We studied the nutritional modes of the orchid Serapias strictiflora and its mycorrhizal fungus Epulorhiza sp. using the differences in carbon isotopic composition (δ¹³C) of C₃ orchid and C₄ maize tissues. We found that if cultivated in substrate lacking any organic compounds, the mycorrhizal extraradical mycelia (δ¹³C = −26.3 ± 0.2 ‰) developed well, despite being fully dependent on nutrition from orchid roots (δ¹³C = −28.6 ± 0.1 ‰). If the mycorrhizal fungus had additional access to and colonized decaying maize roots (δ¹³C = −14.6 ± 0.1 ‰), its isotopic composition (δ¹³C = −21.6 ± 0.4 ‰) reflected a mixture of biotrophy and saprotrophy. No statistically significant differences in δ¹³C of new storage tubers were found between Epulorhiza-associated orchids with (δ¹³C = -28.2 ± 0.1 ‰) and without access to maize roots (δ¹³C = −28.6 ± 0.2 ‰). We conclude that autotrophy is the predominant nutritional mode of mature S. strictiflora plants and that they supply their mycorrhizal fungus with substantial amount of carbon (69 ± 3 % of the fungus demand), even if the fungus feeds saprotrophically.     

<Emphasis Type="Italic">In vitro</Emphasis> regeneration of medicinal plant <Emphasis Type="Italic">Centella asiatica</Emphasis>

This paper describes multiple shoot regeneration from leaf and nodal segments of a medicinally important herb Centella asiatica L. on Murashige and Skoog’s (MS) medium supplemented with a range of growth regulators. The highest number of multiple shoots was observed on MS augmented with 3.0 mg dm⁻³ N⁶-benzylaminopurine (BAP) and 0.05 mg dm⁻³ α-naphthaleneacetic acid (NAA). Leaf explant showed maximum percentage of cultures regenerating shoots (81.6 %), with the highest shoot number (8.3 shoots per explant) and the shoot length (2.1 cm) whereas, nodal explant showed less number of shoots with callus formation at the base cut end. Successive shoot cultures were established by repeatedly sub-culturing the original explant on a fresh medium. Rooting of in vitro raised shoots was best induced on half strength MS supplemented with 0.5 mg dm⁻³ indole-3-butyric acid (IBA) with highest percentage of shoot regenerating roots (76.8 %) with 3–4 roots per shoot. Plantlets were acclimated in Vermi-compost and eventually established in soil. Contents of chlorophyll, total sugars, reducing sugars and proteins were estimated in leaf tissue from both in vivo and in vitro raised plants. Chlorophyll content was higher in in vivo plants, whereas other three components were higher in in vitro plants.     

Different nutrient use strategies of expansive grasses <Emphasis Type="Italic">Calamagrostis epigejos</Emphasis> and <Emphasis Type="Italic">Arrhenatherum elatius</Emphasis>

Enhanced nitrogen (N) levels accelerate expansion of Calamagrostis epigejos and Arrhenatherum elatius, highly aggressive expanders displacing original dry acidophilous grassland vegetation in the Podyjí National Park (Czech Republic). We compared the capability of Calamagrostis and Arrhenatherum under control and N enhanced treatments to (i) accumulate N and phosphorus (P) in plant tissues, (ii) remove N and P from above-ground biomass during senescence and (iii) release N and P from plant material during decomposition of fresh formed litter. In control treatment, significantly higher amounts of total biomass and fresh aboveground litter were observed in Calamagrostis than in Arrhenatherum. Contrariwise, nutrient concentrations were significantly higher (11.6–14.3 mg N g⁻¹ and 2.3 mg P g⁻¹) in Arrhenatherum peak aboveground biomass than in Calamagrostis (8.4–10.3 mg N g⁻¹ and 1.6–1.7 mg P g⁻¹). Substantial differences between species were found in resorption of nutrients, mainly P, at the ends of growing seasons. While P concentrations in Arrhenatherum fresh litter were twice and three times higher (1.6–2.5 mg P g⁻¹) than in Calamagrostis (0.7–0.8 mg P g⁻¹), N concentrations were nearly doubled in Arrhenatherum (13.1–15.6 mg N g⁻¹) in comparison with Calamagrostis (7.4–8.7 mg N g⁻¹). Thus, the nutrients (N and mainly P) were retranslocated from the aboveground biomass of Calamagrostis probably more effectively in comparison with Arrhenatherum at the end of the growing season. On the other hand, Arrhenatherum litter was decomposed faster and consequently nutrient release (mainly N and P) was higher in comparison with Calamagrostis which pointed to different growth and nutrient use strategies of studied grass species.     

<Emphasis Type="Italic">In vitro</Emphasis> bud regeneration of <Emphasis Type="Italic">Carthamus tinctorius</Emphasis> and wild <Emphasis Type="Italic">Carthamus</Emphasis> species from leaf explants and axillary buds

The organogenic competence of leaf explants of eleven Carthamus species including C. tinctorius on Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) + α-naphthaleneacetic acid (NAA) and 6-benzyladenine (BA) + NAA was investigated. Highly prolific adventitious shoot regeneration was observed in C. tinctorius and C. arborescens on both growth regulator combinations and the shoot regeneration frequency was higher on medium supplemented with TDZ + NAA. Nodal culture of nine Carthamus species on media supplemented with BA and kinetin (KIN) individually revealed the superiority of media supplemented with BA over that of KIN in facilitating a higher shoot proliferation index. Proliferating shoots from axillary buds and leaf explants were transferred to medium supplemented with 1.0 mg dm⁻³ KIN or 0.5 mg dm⁻³ BA for shoot elongation. Elongated shoots were rooted on half-strength MS medium supplemented with 1.0 mg dm⁻³ each of indole-butyric acid (IBA) and phloroglucinol. The plantlets thus obtained were hardened and transferred to soil.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

Foliar nutrition of <Emphasis Type="Italic">in vitro</Emphasis>-cultured <Emphasis Type="Italic">Prosopis chilensis</Emphasis> (Molina) Stuntz shoots

Summary Foliar nutrition has been conceived as a possible means of overcoming the recalcitrance of Prosopis chilensis (Molina) Stuntz explants to standard in vitro culture. The foliar uptake of cations (K from 20 gl⁻¹ KNO₃ and Ca from 50 gl⁻¹ CaCl₂), anions (NO₃ from 50 gl⁻¹ KNO₃ and PO₄ from 50 gl⁻¹ NaH₂PO₄), and glucose from a 100 mg l⁻¹ solution studied. All of the nutrients examined were absorbed. The efficacy of foliar nutrition in prolonging the vigor of micropropagated P. chilensis shoot tips was compared with nutrients supplied as a liquid to the base of the stem (liquid) or as an agar-solidified medium (agar). A foliar-feeding apparatus was constructed that employed pressurization of the medium reservoir to drive the medium into the culture vessel with a passive return by a siphoning effect. The medium used was Murashige and Skoog with 30 gl⁻¹ sucrose, 0.1 mgl⁻¹ benzylaminopurine, and 1 mgl⁻¹ indole-3-butyric acid. Over a 9-wk test period it was found that explants cultured by foliar nutrition performed significantly better than those grown on agar for shoot length, nodal production, and leaf retention; and better than liquid MS for node production. There was no significant difference among the three treatments in percentage survival, percentage rooting, or the mean number of roots.     

<Emphasis Type="Italic">In vitro</Emphasis> propagation of <Emphasis Type="Italic">Gentiana dinarica</Emphasis> Beck.

Gentiana dinarica Beck, rare and endangered species of Balkan Dinaric alps, was in vitro propagated (micropropagated) from axillary buds of plants collected at Mt. Tara, Serbia. G. dinarica preferred MS to WPM medium, with optimal shoot multiplication on MS medium with 3% sucrose, 1.0 mg l⁻¹ BA and 0.1 mg l⁻¹ NAA. Rooting was not clearly separated from shoot multiplication since BA did not completely inhibit root initiation. Spontaneous rooting on plant growth regulator-free medium occurred in some 30% of shoot explants. Rooting was stimulated mostly by decreased mineral salt nutrition and a medium with 0.5 MS salts, 2% sucrose and 0.5–1.0 mg l⁻¹ IBA was considered to be optimal for rooting. Rooted plantlets were successfully acclimated and further cultured in peat-based substrate.     

Enhancement of phenylethanoid glycosides biosynthesis in cell cultures of <Emphasis Type="Italic">Cistanche deserticola</Emphasis> by osmotic stress

The effect of osmotic stress on cell growth and phenylethanoid glycosides (PeGs) biosynthesis was investigated in cell suspension cultures of Cistanche deserticola Y. C. Ma, a desert medicinal plant grown in west region of China. Various initial sucrose concentrations significantly affected cell growth and PeGs biosynthesis in the suspension cultures, and the highest dry weight and PeGs accumulation reached 15.9 g l⁻¹-DW and 20.7 mg g⁻¹-DW respectively at the initial osmotic stress of 300 mOsm kg⁻¹ where the sucrose concentration was 175.3 mM. Stoichiometric analysis with different combinations of sucrose and non-metabolic sugar (mannitol) or non-sugar osmotic agents (PEG and NaCl) revealed that osmotic stress itself was an important factor for enhancing PeGs biosynthesis in cell suspension cultures of C. deserticola. The maximum PeGs contents of 26.9 and 23.8 mg g⁻¹-DW were obtained after 21 days at the combinations of 87.6 mM sucrose with 164.7 mM mannitol (303 mOsm kg⁻¹) or 20 mM PEG respectively, which was higher than that of C. deserticola cell cultures grown under an initial sucrose concentration of 175.3 mM after 30 days. The stimulated PeGs accumulation in the cell suspension cultures was correlated to the increase of phenylalanine ammonium lyase (PAL) activity induced by osmotic stress.     

Compatibility of <Emphasis Type="Italic">Piriformospora indica</Emphasis> and <Emphasis Type="Italic">Trichoderma harzianum</Emphasis> as dual inoculants in black pepper (<Emphasis Type="Italic">Piper nigrum</Emphasis> L.)

The compatibility of two biological inoculants, Trichoderma harzianum, a mycoparasitic biological control fungus and Piriformospora indica, a root colonizing plant-growth promoting endophytic fungus was evaluated using tissue cultured black pepper plantlets. We report, for the first time, the ability of P. indica to colonize black pepper, a perennial climber. T. harzianum inhibited the growth of P. indica in an in vitro dual culture plate assay. Simultaneous inoculation with both biological inoculants of tissue cultured black pepper plantlets negatively influenced root colonization by P. indica. However, when P. indica was applied initially followed 30 days later by T. harzianum, there was increased root colonization by the root endophyte P. indica and beneficial effects were found on the growth of the black pepper plants. The present study also showed that the efficacy of inoculation of the two fungal biological agents can be increased by sequential application of P. indica at the hardening stage followed by T. harzianum during transplanting into a soil-sand mixture.