Isolation of micropropagated strawberry endophytic bacteria and assessment of their potential for plant growth promotion

Twenty endophytic bacteria were isolated from the meristematic tissues of three varieties of strawberry cultivated in vitro, and further identified, by FAME profile, into the genera Bacillus and Sphingopyxis. The strains were also characterized according to indole acetic acid production, phosphate solubilization and potential for plant growth promotion. Results showed that 15 strains produced high levels of IAA and all 20 showed potential for solubilizing inorganic phosphate. Plant growth promotion evaluated under greenhouse conditions revealed the ability of the strains to enhance the root number, length and dry weight and also the leaf number, petiole length and dry weight of the aerial portion. Seven Bacillus spp. strains promoted root development and one strain of Sphingopyxis sp. promoted the development of plant shoots. The plant growth promotion showed to be correlated to IAA production and phosphate solubilization. The data also suggested that bacterial effects could potentially be harnessed to promote plant growth during seedling acclimatization in strawberry.     

Isolation and Identification of Bacillus amyloliquefaciens IBFCBF‐1 with Potential for Biological Control of Phytophthora Blight and Growth Promotion of Pepper

In this study, 76 bacterial strains were isolated from the rhizosphere soil of pepper. Of these, 23 bacterial isolates capable of inhibiting Phytophthora capsici growth were selected. Among the antagonistic bacteria, one strain, IBFCBF‐1 showed the strongest antagonistic activity, and was identified as Bacillus amyloliquefaciens based on the results of 16S rRNA gene sequence analysis, physiological and biochemical testing, and morphological characteristics. When tested with a dual‐culture method and with laboratory greenhouse studies, the strain IBFCBF‐1 was found to be a potential biocontrol agent for controlling the plant pathogen, P. capsici. Moreover, it showed high efficiency and broad‐spectrum antifungal properties in vitro. Under greenhouse conditions, IBFCBF‐1 could significantly promote the growth of pepper seedlings, and was able to solubilize phosphate, and produce indole acetic acid (IAA) and ammonia. This study clearly demonstrated that IBFCBF‐1 is a potential candidate exhibiting phytophthora blight‐suppressive and plant growth‐promoting effects on pepper.     

Bacteria isolated from soils of the western Amazon and from rehabilitated bauxite-mining areas have potential as plant growth promoters

Several processes that promote plant growth were investigated in endophytic and symbiotic bacteria isolated from cowpea and siratro nodules and also in bacterial strains recommended for the inoculation of cowpea beans. The processes verified in 31 strains were: antagonism against phytopathogenic fungi, free-living biological nitrogen fixation, solubilization of insoluble phosphates and indole acetic acid (IAA) production. The resistance to antibiotics was also assessed. Sequencing of the partial 16S rRNA gene was performed and the strains were identified as belonging to different genera. Eight strains, including some identified as Burkholderia fungorum, fixed nitrogen in the free-living state. Eighteen strains exhibited potential to solubilize calcium phosphate, and 13 strains could solubilize aluminum phosphate. High levels of IAA production were recorded with l-tryptophan addition for the strain UFLA04-321 (42.3 μg mL^?1). Strains highly efficient in symbiosis with cowpea bean, including strains already approved as inoculants showed the ability to perform other processes that promote plant growth. Besides, these strains exhibited resistance to several antibiotics. The ability of the nitrogen-fixing bacteria to perform other processes and their adaptation to environmental conditions add value to these strains, which could lead to improved inoculants for plant growth and environmental quality.     

Sugarcane Growth Promotion by the Endophytic Bacterium Pantoea agglomerans 33.1

The promotion of sugarcane growth by the endophytic Pantoea agglomerans strain 33.1 was studied under gnotobiotic and greenhouse conditions. The green fluorescent protein (GFP)-tagged strain P. agglomerans 33.1::pNKGFP was monitored in vitro in sugarcane plants by microscopy, reisolation, and quantitative PCR (qPCR). Using qPCR and reisolation 4 and 15 days after inoculation, we observed that GFP-tagged strains reached similar density levels both in the rhizosphere and inside the roots and aerial plant tissues. Microscopic analysis was performed at 5, 10, and 18 days after inoculation. Under greenhouse conditions, P. agglomerans 33.1-inoculated sugarcane plants presented more dry mass 30 days after inoculation. Cross-colonization was confirmed by reisolation of the GFP-tagged strain. These data demonstrate that 33.1::pNKGFP is a superior colonizer of sugarcane due to its ability to colonize a number of different plant parts. The growth promotion observed in colonized plants may be related to the ability of P. agglomerans 33.1 to synthesize indoleacetic acid and solubilize phosphate. Additionally, this strain may trigger chitinase and cellulase production by plant roots, suggesting the induction of a plant defense system. However, levels of indigenous bacterial colonization did not vary between inoculated and noninoculated sugarcane plants under greenhouse conditions, suggesting that the presence of P. agglomerans 33.1 has no effect on these communities. In this study, different techniques were used to monitor 33.1::pNKGFP during sugarcane cross-colonization, and our results suggested that this plant growth promoter could be used with other crops. The interaction between sugarcane and P. agglomerans 33.1 has important benefits that promote the plant''s growth and fitness.     

In vitro desiccation tolerance of the epiphytic Ghost Orchid,Dendrophylax lindenii (Lindl.) Benth x. Rolfe

The Ghost Orchid, Dendrophylax lindenii (Lindl.) Benth x. Rolfe, is a rare and endangered epiphytic orchid native to south Florida and Cuba. The orchid is considered difficult to propagate under greenhouse conditions, requiring high humidity and low air movement. In contrast, the orchid’s native habitat seasonally dries out with decreased precipitation and humidity. This suggests some level of desiccation tolerance. Ghost Orchid plants were assessed for potential desiccation tolerance and ability to recover from desiccation stress under in vitro conditions. In vitro-derived plants were placed into sterile baby food jars and transferred to chambers maintained at 10% relative humidity, which is extremely low compared to relative humidity levels (40–100%) recorded under natural field conditions. Plants were removed every week for 4 wk and recovered on P723 medium supplemented with banana powder for 4 wk. Data were collected at the initiation of the experiment, after the desiccation periods, and after 4 wk in vitro recovery. Ghost Orchid plants demonstrated extremely high desiccation tolerance. Even after 4 wk desiccation, plant survival was observed at 79.2% after recovery. Desiccated plants exhibited significant decreases in tissue water potential (− 18.44 MPa), fresh weight (65.5% loss), and water content (14.2%); however, high plant survival was still observed under these conditions similar to poikilohydric plants. Overall, the Ghost Orchid demonstrated high desiccation tolerance, which should be considered for future greenhouse culture and for its application in the direct field establishment of in vitro-derived plants without greenhouse acclimatization.

     

Effect of co-inoculation of methylotrophic Methylobacterium oryzae with Azospirillum brasilense and Burkholderia pyrrocinia on the growth and nutrient uptake of tomato, red pepper and rice

The present greenhouse study was undertaken to evaluate the effects of co-inoculating methylotrophic Methylobacterium oryzae CBMB20 along with nitrogen-fixing Azospirillum brasilense CW903 or a phosphate solubilizing bacterium Burkholderia pyrrocinia CBPB-HOD on the growth and nutrient uptake of tomato, red pepper and rice. Seed inoculation and soil/foliar application of the bacterial strains alone or under dual inoculation increased the plant growth in terms of shoot or root length and increased the nutrient uptake in the plants studied compared to uninoculated control plants. Co-inoculation of M. oryzae CBMB20 with A. brasilense CW903 or B. pyrrocinia CBPB-HOD improved the N and P concentration of plants, while the results varied among the plant species tested. Also, co-inoculation of the bacterial strains increased the activity of nitrogenase, urease and phosphatase enzymes in soil when compared to uninoculated control or individual inoculations. Though the inoculation effects were analyzed at an early stage of plant growth, the results conclusively suggest that M. oryzae being compatible with other microorganisms in the rhizosphere can potentially be used as individual inoculant or co-inoculated with other plant growth promoting bacteria to increase the production in sustainable agricultural systems.     

Synthesis of Japanese Boletus edulis ectomycorrhizae with Japanese red pine

Boletus edulis is a well-known ectomycorrhizal mushroom. Although cultivation has been widely attempted, no artificial fruiting has been achieved owing to difficulties associated with mycorrhizal synthesis and acclimatization in fields. We collected fifteen B. edulis basidiomata samples from locations in Japan and identified them microscopically and by phylogenetic analysis of their nuclear ribosomal internal transcribed spacer (ITS) regions. Pure culture isolates of B. edulis were established efficiently on malt extract agar medium, and one isolate, EN-63, was inoculated to axenic Pinus densiflora seedlings in vitro. Brownish ectomycorrhizal tips were observed on the pine lateral roots within four months of inoculation. Ten pine seedlings that formed ectomycorrhizae were acclimatized under laboratory and greenhouse conditions. At four months after transplant, mycorrhizal colonization by B. edulis was observed on newly grown root tips under laboratory conditions, but no B. edulis ectomycorrhiza survived under greenhouse conditions. These results suggest that B. edulis ectomycorrhizae synthesized in vitro with P. densiflora requires additional steps for acclimatization to greenhouse conditions.     

Phosphate solubilizing rhizobacteria as alternative of chemical fertilizer for growth and yield of Triticum aestivum (Var. Galaxy 2013)

AimThe presence of Phosphorus as a macronutrient in soil is necessary for plant growth and its deficiency restricts crop yield. Therefore, the aim of current study is to isolate promising rhizospheric phosphate solubilizing bacteria presenting with plant growth promoting (PGP) traits and their utilization as biofertilizers to improve Triticum aestivum (Var. Galaxy 2013) growth and nutrition.MethodOut of 30 isolates obtained from rhizosphere of various plants of different regions, 10 best PSRB strains (WumS-3, WumS-4, WumS-5, WumS-11, WumS-12, WumS-21, WumS-24, WumS-25, WumS-26 and WumS-28) were selected based on their high P solubilization and good PGP (auxin, psiderphore, HCN, Nitrogen fixation) activities. Triticum aestivum (Var. Galaxy 2013) was used as an experimental crop under laboratory and field conditions.ResultsIn this study, P solubilization capacity of selected strains were found 4–7 solubilization index on agar plate and 30–246 µg/ml in liquid broth respectively. The optimum conditions for phosphate solubilization under in vitro condition were found 35 °C at pH 7, glucose as good carbon source and ammonium nitrate as a good nitrogen source. Furthermore, the selected strains had the ability to produces phytohormones (indole acetic acid), siderophore, ammonia and Hydrogen Cyanide. Finally, PSRB inoculum showed significant (p < 0.05) increase (50%–80%) in seed germination while 10–90% increase in root length and shoot length was found as compared to control in laboratory condition. Under natural conditions, 40–80% increase in seed germination while 5–34.8% increase in shoot length and 5–96% increase in seed weight was also observed.ConclusionIsolated strains are promising PSRB that enhance plant growth and this research is a base for recommending the use of these bacterial strains for biofertilizer, as an alternative of chemical fertilizer, for Triticum aestivum L. production.     

Assessing European Egg Parasitoids as a Mean of Controlling the Invasive South American Tomato Pinworm Tuta absoluta

The South American tomato pinworm (Tuta absoluta) has recently invaded Europe and is rapidly spreading in the Afro-Eurasian continent where it is becoming a major pest on tomato crops. Laboratory tests were undertaken to evaluate the potential of 29 European strains of Trichogramma parasitoids to control T. absoluta. In addition to the host itself, the host plant (tomato) was used during the laboratory tests in order to increase the chance of selecting the best parasitoid strains. Trichogramma females were placed with T. absoluta eggs on a tomato leaflet in tubes. We compared the parasitism of T. absoluta by the various Trichogramma species tested to the Trichogramma species currently commercially available for the pest control in Europe, i.e. Trichogramma achaeae. Thereafter, the more promising strains were tested on a larger scale, in mesocosm (i.e. cages in greenhouses) and in greenhouse compartments to evaluate efficiency of laboratory selected strains under cropping conditions. The most efficient strain from the laboratory screening trials did not perform as efficiently under the greenhouse conditions. We discuss differences in parasitism levels among species and strains and among the different scales tested in the experiments, as well as implications of these results for further screening for biocontrol agents.     

Plant growth promoting bacteria from Crocus sativus rhizosphere

Present study deals with the isolation of rhizobacteria and selection of plant growth promoting bacteria from Crocus sativus (Saffron) rhizosphere during its flowering period (October–November). Bacterial load was compared between rhizosphere and bulk soil by counting CFU/gm of roots and soil respectively, and was found to be ~40 times more in rhizosphere. In total 100 bacterial isolates were selected randomly from rhizosphere and bulk soil (50 each) and screened for in-vitro and in vivo plant growth promoting properties. The randomly isolated bacteria were identified by microscopy, biochemical tests and sequence homology of V1–V3 region of 16S rRNA gene. Polyphasic identification categorized Saffron rhizobacteria and bulk soil bacteria into sixteen different bacterial species with Bacillus aryabhattai (WRF5-rhizosphere; WBF3, WBF4A and WBF4B-bulk soil) common to both rhizosphere as well as bulk soil. Pseudomonas sp. in rhizosphere and Bacillus and Brevibacterium sp. in the bulk soil were the predominant genera respectively. The isolated rhizobacteria were screened for plant growth promotion activity like phosphate solubilization, siderophore and indole acetic acid production. 50 % produced siderophore and 33 % were able to solubilize phosphate whereas all the rhizobacterial isolates produced indole acetic acid. The six potential PGPR showing in vitro activities were used in pot trial to check their efficacy in vivo. These bacteria consortia demonstrated in vivo PGP activity and can be used as PGPR in Saffron as biofertilizers.This is the first report on the isolation of rhizobacteria from the Saffron rhizosphere, screening for plant growth promoting bacteria and their effect on the growth of Saffron plant.     

Asymbiotic seed germination and in vitro seedling development of <Emphasis Type="Italic">Cyrtopodium punctatum</Emphasis>: a propagation protocol for an endangered Florida native orchid

Cyrtopodium punctatum Lindley is an endangered epiphytic orchid restricted in the United States to southern Florida. Due to its ornamental value, the species was extensively collected from the wild during the past 100 years. Today, only a few plants remain in protected areas. As part of a conservation plan for the species, procedures for asymbiotic seed germination were developed. Five asymbiotic orchid seed germination media (PhytoTechnology Orchid Seed Sowing Medium, Knudson C, Malmgren Modified Terrestrial Orchid Medium, Vacin &; Went Modified Orchid Medium, and ½-strengh Murashige &; Skoog) were examined for their effectiveness in promoting seed germination and protocorm development under a 16/8 h L/D photoperiod and dark (0/24 h L/D). The influence of photoperiod on growth and development was also examined. Seeds were germinated under a 16/8 h, 12/12 h, 8/16 h L/D photoperiod, at 25 ± 3°C and allowed to develop in vitro for 10 weeks. After 10 weeks, developing seedlings were transferred to Sigma Phytatrays and returned to their assigned photoperiod treatments for continued seedling development for an additional 15 weeks. Highest germination occurred in 0/24 h L/D on PhytoTechnology Orchid Seed Sowing Medium and seedlings displayed more advanced development when cultured under 16/8 h L/D photoperiod after 15 weeks in Phytatrays. Thirty-five week old seedlings potted in coconut husk growing medium exhibited 90% survival following 5 weeks acclimatization to greenhouse conditions. This asymbiotic seed germination protocol for C. punctatum will facilitate future reintroduction projects involving this endangered species.     

Carob trees (Ceratonia siliqua L.) regenerated in vitro can acclimatize successfully to match the field performance of seed-derived plants

The use of in vitro regenerated plants in forestry and orchard depends ultimately on the development of efficient transplantation protocols, ensuring high survival rates and successful establishment under field conditions. We tested the performance of micropropagated carob trees (Ceratonia siliqua L.) throughout the acclimatization process in terms of survival, growth and physiological traits, including field comparisons with seed-derived and mother plants. The field trial was 100?% successful, i.e. we found no major differences between micropropagated, seed-derived and mother plants in terms of growth rate, height, number of leaves, photosynthetic efficiency, chlorophyll fluorescence, chlorophyll content and soluble protein content, although these parameters changed significantly during acclimatization. Stomatal conductance (g _s) was reduced by fourfold when plants were transferred from in vitro culture to the growth chamber, thus preventing uncontrolled wilting. The photosynthetic rate (P _N) was relatively low in vitro, in the growth chamber and the greenhouse, but increased to match seed-derived and mother plants in the field. The chlorophyll a/b ratio in leaves from in vitro and growth chamber plants was typical of shade plants (2.1) but became more characteristic of sun plants in the subsequent acclimatization stages (3.1–3.5). The maximum efficiency of photosystem II (F _v/F _m) remained mostly constant at?~0.80 throughout acclimatization, as is typical for healthy, non-stressed plants. We conclude that our micropropagation and acclimatization protocols provide a suitable alternative to traditional mass propagation techniques for the carob tree.     

Inherently slow growth in two Caribbean epiphytic species: A demographic approach

Abstract. In relation to the drought‐prone and nutrient‐poor habitat, vascular epiphytes are routinely referred to as inherently slow‐growing plants, although actual evidence is rare. To test this notion we measured in situ growth of the understorey orchid Aspasia principissa and the tank bromeliad Vriesea sanguinolenta, and, for the latter species, also the growth under favourable conditions in the greenhouse. Using growth analysis we show: (1) that in an intraspecific comparison, small to intermediate individuals yield the highest relative growth rates (RGR) in situ: A. principissa: 1.6 10–³ d^‐1; V. sanguinolenta: 3.3 10–³ d^‐1; (2) that the bromeliad reaches maximum size after ca. 15 yr, while the orchid needs at least 20 yr; and (3) small V. sanguinolenta plants exhibit a highly plastic growth response to favourable conditions in the greenhouse, reaching an almost 10‐fold increase in RGR. In spite of a substantial increase in growth under more favourable conditions, our results are consistent with the notion that epiphytes are inherently slow growing organisms.     

In Vitro and In Vivo Plant Growth Promoting Activities and DNA Fingerprinting of Antagonistic Endophytic Actinomycetes Associates with Medicinal Plants

Endophytic actinomycetes have shown unique plant growth promoting as well as antagonistic activity against fungal phytopathogens. In the present study forty-two endophytic actinomycetes recovered from medicinal plants were evaluated for their antagonistic potential and plant growth-promoting abilities. Twenty-two isolates which showed the inhibitory activity against at least one pathogen were subsequently tested for their plant-growth promoting activities and were compared genotypically using DNA based fingerprinting, including enterobacterial repetitive intergenic consensus (ERIC) and BOX repetitive elements. Genetic relatedness based on both ERIC and BOX-PCR generates specific patterns corresponding to particular genotypes. Exponentially grown antagonistic isolates were used to evaluate phosphate solubilization, siderophores, HCN, ammonia, chitinase, indole-3-acetic acid production, as well as antifungal activities. Out of 22 isolates, the amount of indole-3-acetic acid (IAA) ranging between 10–32 μg/ml was produced by 20 isolates and all isolates were positive for ammonia production ranging between 5.2 to 54 mg/ml. Among 22 isolates tested, the amount of hydroxamate-type siderophores were produced by 16 isolates ranging between 5.2 to 36.4 μg/ml, while catechols-type siderophores produced by 5 isolates ranging from 3.2 to 5.4 μg/ml. Fourteen isolates showed the solubilisation of inorganic phosphorous ranging from 3.2 to 32.6 mg/100ml. Chitinase and HCN production was shown by 19 and 15 different isolates, respectively. In addition, genes of indole acetic acid (iaaM) and chitinase (chiC) were successively amplified from 20 and 19 isolates respectively. The two potential strains Streptomyces sp. (BPSAC34) and Leifsonia xyli (BPSAC24) were tested in vivo and improved a range of growth parameters in chilli (Capsicum annuum L.) under greenhouse conditions. This study is the first published report that actinomycetes can be isolated as endophytes from within these plants and were shown to have antagonistic and plant growth promoting abilities. These results clearly suggest the possibility of using endophytic actinomycetes as bioinoculant for plant growth promotion, nutrient mobilization or as biocontrol agent against fungal phytopathogens for sustainable agriculture.     

A low-cost alternative membrane system that promotes growth in nodal cultures of Brazilian ginseng [Pfaffia glomerata (Spreng.) Pedersen]

In vitro propagated plants under conditions of low gas exchange generally show morphological and physiological anomalies that lead to high mortality rates during ex vitro acclimatization. The use of gas-permeable membranes increases natural ventilation in culture vessels, photosynthesis and growth rates. However, commercial membranes are expensive, which limits their application. In this study, low-cost, simple to manufacture, alternative membranes were developed to promote gas exchange in jars used for in vitro plant tissue culture. The membranes were developed using polytetrafluoroethylene film and two or three layers of microporous tape (Missner & Missner^?), and were designed to increase the growth of nodal cultures of Pfaffia glomerata (Brazilian ginseng). Conditions that provided higher gas exchange led to an increase in plant growth and content of photosynthetic pigments compared to a closed system without a gas-permeable membrane. The alternative membranes showed similar results for water vapor loss rate and photosynthetic pigments when compared to a commercial membrane. The alternative membranes were also an efficient barrier against contamination and remained intact after being autoclaved multiple times. Among the membranes tested, the traits of the P. glomerata in vitro-derived plants were similar when propagated using the alternative membrane with three layers of microporous tape or the commercial membrane. However, the alternative membrane has a unit cost that is ten times lower than the commercial membrane.     

Study of the effects of foliar application of ABA during acclimatization

This study aims to elucidate the effects of abscisic acid (ABA) foliar application on plant hardening during ex vitro acclimatization using a forest model species Ulmus minor L. Plant leaves were sprayed with ABA (0, 50 and 100 μM) immediately after ex vitro transfer and twice a week for the first 3 weeks of acclimatization. After this period, parameters related with photosynthesis, hormone levels and oxidative stress were measured to assess plant performance. The results demonstrated that ABA foliar application alleviates the negative shock of ex vitro acclimatization since it reduces the water loss through transpiration, relieving the risk of wilting. Moreover, ABA promoted net CO₂ assimilation rate (P _N ) and plant dry mater accumulation. ABA treatment increased the antioxidant battery during acclimatization, with more effective results at the concentration of 50 μM ABA. Also, flow cytometry data support that cytosolic compounds, which may increase in response to 50 μM ABA, could also protect DNA from oxidative damage. We propose here that ABA foliar application (immediately after ex vitro transfer), by preventing water loss, enhancing photosynthesis efficiency and the activity of antioxidant enzymes, improves the plants hardening and ability to deal with the ex vitro stresses.     

Isolation and characterization of plant growth promoting endophytic diazotrophic bacteria from Korean rice cultivars

We have isolated 576 endophytic bacteria from the leaves, stems, and roots of 10 rice cultivars and identified 12 of them as diazotrophic bacteria using a specific primer set of nif gene. Through 16S rDNA sequence analysis, nifH genes were confirmed in the two species of Penibacillus, three species of Microbacterium, three Bacillus species, and four species of Klebsiella. Rice seeds treated with these plant growth-promoting bacteria (PGPB) showed improved plant growth, increased height and dry weight and antagonistic effects against fungal pathogens. In addition, auxin and siderophore producing ability, and phosphate solubilizing activity were studied for the possible mechanisms of plant growth promotion. Among 12 isolates tested, 10 strains have shown higher auxin producing activity, 6 isolates were confirmed as strains with high siderophore producing activity while 4 isolates turned out to have high phosphate-solubilizing activity. These results strongly suggest that the endophytic diazotrophic bacteria characterized in this study could be successfully used to promote plant growth and inducing fungal resistance in plants.     

Role of halotolerant phosphate‐solubilising bacteria on growth promotion of peanut (Arachis hypogaea) under saline soil

Soil salinity is a major abiotic stress that limits plant growth, and inoculating plant growth‐promoting rhizobacteria is a well‐known strategy to reduce stressors under adverse soil conditions. This study was conducted to assess the effect of halotolerant phosphate‐solubilising bacteria (PSB) on protecting peanut against salt stress. Four candidate strains: Bacillus megaterium (YM13), Enterobacter sp. (YM14), Providencia rettgeri (TPM23) and Ensifer adhaeren (TPMX5) showed strong tolerance to NaCl and high phosphate‐solubilising ability even at a NaCl concentration of 1.4 M. In addition, all four strains demonstrated variable levels of phosphate solubilisation activity in the presence of various carbon and nitrogen sources, indicating high phosphate‐solubilising efficacy. Germination and radicle length of peanut seedlings increased with inoculation of PSB under both control and saline conditions. Statistically significant increase in the root length (range: 25.71–49%), stem length (19–48%), number of leaves (12.5–37%) and root/shoot biomass were observed. This could be attributed to plant hormones (i.e., indole acetic acid [IAA], abscisic acid [ABA] and gibberellic acid [GA3]) and successful root colonisation by bacterial inoculants. Root colonisation was positively correlated to plant growth and shown to be influenced by soil conditions. In addition, the PSB also improved the levels of available P in soil. The most pronounced beneficial effect on the growth of peanut plants and soil available P content was observed in the inoculation of the PSB isolates with Ca₃(PO₄)₂ addition. This is the first report that describes Providencia rettgeri as a plant growth‐promoting bacterium that may be utilised to alleviate the negative effects of salt stress on peanut plants. This bacterial species may thus be potentially used as a biofertilizer for sustaining the growth of peanut in salt‐stressed soil and in mitigating soil stress conditions.     

Establishing clones of Veratrum californicum,a native medicinal species,for micropropagation

Factors affecting the micropropagation of Veratrum californicum, a slow-growing species that is a potentially valuable source of cyclopamine, were investigated. Sterile cultures were initiated on modified Murashige and Skoog medium, and clones from individual donor plants were assigned to experimental conditions when approximately 100 shoots of each clone were available. The effects of temperature, light quality, and plant growth regulators on multiplication and survival were assessed. Four clones from which large greenhouse populations were obtained were selected for in-depth analysis. When shoots were cultured at 10°C and 16°C, multiplication ratios consistently >1 were observed from three of four clones and two of four clones, respectively, during the five-subculture cycles. None of the clones stably increased when cultured at 24°C, and plants from this treatment did not survive acclimatization in the greenhouse. Only one clone showed increased multiplication ratios in response to plant growth regulator treatments, with maximum multiplication when shoots were cultured with 9 μM benzyladenine and 0.5 μM naphthaleneacetic acid. Light quality in the laboratory did not affect multiplication ratio but did affect subsequent greenhouse survival. The size of plants derived from culture was most often equivalent (65% of 1,271) to 3-yr-old seed-derived plants. Although the growth of clones during acclimatization differed, plants derived from cultures incubated at 16°C had the best rates of overall greenhouse survival. Temperature and light treatments in vitro critical to long-term plant survival were demonstrated and will assist the establishment of a mass propagation system for V. californicum.