High Levels Of Germination Research Articles

Germination ecology and growth phenology of cowvine (Ipomoea lonchophylla) as influenced by environmental parameters

AbstractCowvine (Ipomoea lonchophylla J.M. Black) is a native and widely spread summer broadleaf weed in Australia. It contains glycoresins, which are toxic to livestock. However, limited information is available on seed germination ecology and growth phenology of this species. A series of experiments were conducted to determine the response of I. lonchophylla to different environmental conditions. Results showed that the primary dormancy exhibited by I. lonchophylla is due to the physical impediment of the hard seed coat. The seed germination percentage was the highest at the constant temperature of 27 C and alternating temperatures of 35/25 C. Germination of I. lonchophylla was not stimulated by light, suggesting that this species is non-photoblastic. Ipomoea lonchophylla germination was intolerant of a medium to high level of salt stress, and germination was completely inhibited at 250 mM NaCl. The emergence of I. lonchophylla was not restricted by seeding depth up to 8 cm, but only 5% emergence was recorded when seeds were planted at a 16-cm depth. The germination percentage was also drastically reduced by 90% to 100% after exposure to either 3 mo in silage, 48-h digestion in steers, or silage plus digestion treatments. The growth and reproductive phenology of I. lonchophylla was affected by emergence time. Plants that emerged in late spring (November 15) were able to produce more berries per plant than those that emerged in midsummer (January 15) in southern New South Wales. Information gained in our study concerning high soil salinity, ensiling, and digestion will help to develop more sustainable and effective integrated weed management strategies for controlling and reducing the spread of this weed.

Treatment with insecticide and fungicide on the physiological quality of lentil seeds.

The low Brazilian productive index and the high demand have aroused interest in the cultivation of lentils, however the legume is little known and needs further studies. The objective of this study was to analyze and identify the effects of treatments with insecticides and fungicides on the physiological quality of lentil seeds, CA-1512 strain. The experiments were conducted in the seed laboratory in a completely randomized design with seven treatments and four replicates. Seed treatment with Thiophanate-methyl; Fluazinam® (180 ml) + Pyraclostrobin; Thiophanate-methyl; Fipronil® (150 ml) promoted higher levels of germination under accelerated aging, lower number of abnormal seedlings and longer lengths of shoot and radicle for the emergence in paper. Treatment with Carboxin; Thiram® (250 ml) + Imidacloprid® (150 ml) allowed a higher value in the first count of germination in sand, lower number of dead seeds under accelerated aging and longer root length, in the emergence in sand. Shoot length in the emergence in sand increased after seed treatment with Metalaxyl-M; Fludioxonil® (75 ml) + Pyraclostrobin; Thiophanate-methyl; Fipronil® (150 ml). Treatments with fungicides and insecticides considerably improved the physiological properties of the seeds, thus being able to guarantee greater phytosanitary qualities in the field, generating healthier seedlings and with protection against possible pests and diseases, and consequently guaranteeing greater productivity.

In vitro regeneration of Senna alata (L.) Robx. (Fabaceae): An ancient medicinal plant

Senna alata, a legume of medicinal importance, presents antifungal and antibacterial antioxidant activity. The main limitation is the presence of seeds with integument dormancy within natural populations. This study aimed to establish an efficient protocol for the in vitro propagation and acclimatization of seedlings. The seeds were disinfected with different concentrations of NaOCl; for the regeneration of the shoots, cotyledon segments were inoculated in MS and WPM medium, supplemented with plant growth regulators, auxins (1-NAA and 2,4-D), interaction with cytokinins (TDZ and BAP), and activated carbon. The seedlings were acclimated in nursery conditions using vermiculite, sand, and soil substrates. Using NaOCl (0.5%) for 30 min resulted in 98.0% disinfected seeds while 95.4% of scarified seeds germinated. High node and shoots numbers (8.3 ± 0.4; 12.6 ± 0.8, respectively) were obtained when using the MS medium containing 1-NAA + TDZ (0.1 + 1.0 mg L-1), while the use of MS medium supplemented with 2,4-D + TDZ (0.1 + 1.0 mg L-1) resulted in longer shoots (6.0 ± 0.5 cm). Using WPM medium with IBA + BAP (1.0 + 0.1 mg L-1) improved rooting (14.0 cm ± 1.3) and the number of roots (5.0 ± 0.0) in the treated plants. The vermiculite substrate with sand promoted 81.5% of acclimatized plants. This study allowed obtaining high levels of germination, regeneration in vitro and acclimatization of S. alata.

Efficient Secretion of Murine IL-2 From an Attenuated Strain of Clostridium sporogenes, a Novel Delivery Vehicle for Cancer Immunotherapy.

Despite a history dating back to the 1800s, using Clostridium bacteria to treat cancer has not advanced beyond the observation that they can colonise and partially destroy solid tumours. Progress has been hampered by their inability to eradicate the viable portion of tumours, and an instinctive anxiety around injecting patients with a bacterium whose close relatives cause tetanus and botulism. However, recent advances in techniques to genetically engineer Clostridium species gives cause to revisit this concept. This paper illustrates these developments through the attenuation of C. sporogenes to enhance its clinical safety, and through the expression and secretion of an immunotherapeutic. An 8.6 kb sequence, corresponding to a haemolysin operon, was deleted from the genome and replaced with a short non-coding sequence. The resultant phenotype of this strain, named C. sporogenes-NT, showed a reduction of haemolysis to levels similar to the probiotic strain, C. butyricum M588. Comparison to the parental strain showed no change in growth or sporulation. Following injection of tumour-bearing mice with purified spores of the attenuated strain, high levels of germination were detected in all tumours. Very low levels of spores and vegetative cells were detected in the spleen and lymph nodes. The new strain was transformed with four different murine IL-2-expressing plasmids, differentiated by promoter and signal peptide sequences. Biologically active mIL-2, recovered from the extracellular fraction of bacterial cultures, was shown to stimulate proliferation of T cells. With this investigation we propose a new, safer candidate for intratumoral delivery of cancer immunotherapeutics.

Improving the operation of a drum grain dryer with justification of the low-temperature mode parameters

The proposed engineering and technological solutions are aimed at improving the operation of a low-temperature drum grain dryer by using a combined mechanical vibration exciter, with the further justification of low-temperature regime parameters. Existing vibration technologies imply the high-temperature drying of food grains with further utilization of spent heat carrier for reheating it. In this case, the high-temperature drying of cereals and seed crops (grains, onions, etc.) does not make it possible to maintain high germination, emphasizing the need for low-temperature treatment, which, in this case, reduces the efficiency of spent heat carrier. Therefore, to improve the efficiency of the drying process and technology, it has been proposed to employ vibration low-frequency technologies that ensure seed quality. The trajectory and the kinetic energy of the drum container with a combined mechanical vibration exciter have been calculated for the improved vibratory drum grain dryer. The rational vibration intensity for seed drying providing the following kinetic characteristics has been established: vibration speed, to 0.03 m/s; vibration acceleration, 30 m/s 2 ; vibration intensity, 2.6 m 2 /s 2 ; at amplitude vibration not exceeding 2 mm. It has been also established that the intense warming of a barley layer occurs at a temperature of 50 °C and a humidity of 13.5 %, the final temperature is 42.4 °C, and, at 40 °C, is, accordingly, 35.4 °C. This has confirmed that the low-temperature drying of barley seeds of the variety Stalker (Ukraine) takes place during the periods of constant and falling drying speed characterized by a high level of germination (95...93 %)

Efekat hemijske desikacije na klijavost i čuvanje semena suncokreta

The objective was to assess the effect of chemical desiccation on seed germination and storability in three sunflower female inbred lines and to point at possible indicators for optimal performance time. Desiccation was performed with Diquat (2 l ha-1) applied at 7-day intervals from the end of flowering to harvest maturity. There were 6 treatments and control (untreated). Germination was tested 2 and 21 months after harvest. The highest germination was recorded when the desiccation was performed 35 days after flowering. Results showed that optimal moment for applying chemical desiccation when there is no negative effect on the seed germination is specific for each sunflower genotype. Seeds with a high level of germination (>90%) can be stored for 19 months without significant loss in germination, namely, it was not negatively affected by chemical desiccation. Seed moisture and growing degree-days can be used as reliable indicators for optimal desiccation time.

Neem oil increases the persistence of the entomopathogenic fungus Metarhizium anisopliae for the control of Aedes aegypti (Diptera: Culicidae) larvae

BackgroundThe entomopathogenic fungus Metarhizium anisopliae is a candidate for the integrated management of the disease vector mosquito Aedes aegypti. Metarhizium anisopliae is pathogenic and virulent against Ae. aegypti larvae; however, its half-life is short without employing adjuvants. Here, we investigated the use of neem oil to increase virulence and persistence of the fungus under laboratory and simulated field conditions.MethodsNeem was mixed with M. anisopliae and added to recipients. Larvae were then placed in recipients at 5-day intervals for up to 50 days. Survival rates were evaluated 7 days after exposing larvae to each treatment. The effect of neem on conidial germination following exposure to ultraviolet radiation was evaluated under laboratory conditions. Statistical tests were carried out using ANOVA and regression analysis.ResultsLaboratory bioassays showed that the fungus alone reduced survival to 30% when larvae were exposed to the treatment as soon as the suspension had been prepared (time zero). A mixture of fungus + neem resulted in 11% survival at time zero. The combination of fungus + neem significantly reduced larval survival rates even when suspensions had been maintained for up to 45 days before adding larvae. For simulated-field experiments 1% neem was used, even though this concentration is insecticidal, resulting in 20% survival at time zero. However, this toxic effect was reduced over time. When used alone under simulated-field conditions the fungus rapidly lost virulence. The formulation fungus + neem effectively maintained fungal virulence, with larval survival rates significantly reduced for up to 45 days after preparation of the suspensions. The effective half-life of the fungus or neem when used separately was 6 and 13 days, respectively. The half-life of fungus formulated in 1% neem was 34 days. Conidia suspended in neem maintained high levels of germination even following a 2-h exposure to ultraviolet radiation.ConclusionsA combination of the entomopathogenic fungus M. anisopliae with neem oil effectively increases the half-life and virulence of the fungus when tested against Ae. aegypti larvae, even under simulated field conditions. Neem oil also protected the fungus from the damaging effects of ultraviolet radiation.

The effect of various after-ripening temperature regimens on the germination behaviour of Ambrosia artemisiifolia

Ambrosia artemisiifolia (common ragweed) is an annual weed known to infest crops and disturbed areas, and cause human pollinosis. To examine the effect of different thermal regimens on seed germination, during seed after-ripening, a study was conducted for two years. In 2012 (year 1) and 2013 (year 2), ragweed seeds collected from a single wheat stubble field were divided and stored under one of the five temperature regimens: constant −20 °C, +5 °C, room temperature (18 °C), +25 °C, and field conditions. Germination tests were performed every 15 days of storage from day 17 to day 213. Storage conditions showed a strong influence on ragweed seed germination duration. At constant low temperatures (−20 °C), seed behaved in an opposite manner depending on the year; in year 1 germination was inhibited while it was stimulated in year 2. At higher temperatures, seed dormancy was unbroken due to a lack of exposure to low temperatures. Under field conditions, seeds reached a high level of germination after a few days of storage due to temperature fluctuations. The different behaviours of ragweed seeds shown at constant temperatures revealed different dormancy depths in the two years that might be due to population variability and maternal environment differences during seed maturation.

DNA Replication during Seed Germination, Deterioration, and Its Relation to Vigor in Alfalfa and White Clover

Differences in seed vigor that arise due to seed aging, even in seeds with high levels of germination, can result in differences in field emergence. Current test methods to identify vigor differences in commercial seed lots with high standard germination take several days to complete. DNA replication, measured using flow cytometry as the ratio of 4C/2C DNA, during early germination of unaged and aged seeds of alfalfa (Medicago sativa L.) and white clover (Trifolium repens L.) was examined as a potential rapid and accurate method of assessing seed vigor. The relationship between 4C/2C DNA during germination of 10 commercial seed lots of both species and their field emergence was also determined. Most of the nuclei in dry mature seeds were arrested in the 2C DNA stage, with only a small proportion of 4C DNA nuclei. The proportion of 4C nuclei increased during germination of seeds, although the increase was lower and delayed in aged seeds with slower and lower germination. The 4C/2C ratio after 24‐h imbibition predicted the field emergence of all seed lots, but particularly of the lots with high standard germination (SG) (SG ≥ 75%, alfalfa, R2 = 0.815; SG ≥ 80%, white clover, R2 = 0.771; P < 0.01). Thus, DNA replication marks an advancement of germination in alfalfa and white clover seed, and the 4C/2C DNA ratio after 24‐h imbibition has potential to predict the seed vigor of both species.

More butenolides from plant-derived smoke with germination inhibitory activity against karrikinolide

Plant-derived smoke is known to play an important role in promoting germination in the post-fire environment, thereby stimulating regeneration of vegetation from the soil seed bank following a fire. Several compounds with germination promotory activity have been identified in recent years, with one such compound, 3-methyl-2H-furo[2,3-c]pyran-2-one (karrikinolide; KAR1) known to be highly active in promoting germination of many plant species at low concentrations. In contrast, a related compound, (5RS)-3,4,5-trimethylfuran-2(5H)-one, also found in plant-derived smoke, was shown to have an antagonistic effect against KAR1, resulting in lowered germination levels when tested on light-sensitive ‘Grand Rapids’ lettuce seeds which usually have high germination levels when treated with KAR1. Considering the combination of such types of compounds in smoke, there is the possibility of smoke having a dual regulatory mechanism to control post-fire seedling emergence. Here we report the identification of two structurally related butenolides from plant-derived smoke with germination inhibitory activity, namely 5,5-dimethylfuran-2(5H)-one and (5RS)-5-ethylfuran-2(5H)-one. In the germination test system using light-sensitive ‘Grand Rapids’ lettuce seeds, the 5,5-dimethylfuran-2(5H)-one showed similar levels of activity against KAR1 as previously observed with the (5RS)-3,4,5-trimethylfuran-2(5H)-one, whereas the (5RS)-5-ethylfuran-2(5H)-one was approximately 100 times less active.

Effect of light, gibberellic acid and nitrogen source on germination of eight taxa from dissapearing European temperate forest, Potentillo albae-Quercetum

Little is known about how light affects seed germination and revegetation of species of thermophilous oak forest. To reveal this relationship effects of white, red, far-red irradiations and dark incubation on germination of eight Potentillo albae-Quercetum taxa were examined. Attempts were also made to evaluate the influence of gibberellic acid and different nitrogen sources on the germination characteristics. Interaction between light and nitrogen was also studied. Freshly matured seeds of all taxa germinated very poorly, indicating presence of primary dormancy. Germination rates increased after wet-stratification treatment and were low in darkness. The highest concentration of the nitrogenous solutions that resulted in high germination level was 10 mM, whereas higher concentrations had a negative effect. Nitrate had the strongest influence which can be proved by a ‘gap detection’ mechanism for gaps in the vegetation. Far-red and red irradiation showed antagonistic effect on seed germination. There was a decrease in germination when far-red was followed by red and an improvement when red was followed by far-red treatment. Under red light, gibberellic acid enhanced germination of positively photoblastic taxa. It was concluded that light factor, associated with vegetation gaps, was the most important signal stimulating germination of the studied taxa.

Crop resistance to parasites.

Crop Science The parasitic weed Striga infests a large proportion of cereal crops in Africa and parts of Asia, which has a devastating effect on farming, particularly in subsistence communities. An obligate parasite, Striga requires a chemical signal from the host plant to germinate, offering an avenue for genetic improvement of host crops. Gobena et al. studied the difference between sorghum strains associated with high and low Striga germination levels and mapped a mutation associated with low Striga germination ( LGS1 ) to a previously uncharacterized sulfotransferase gene. Deletion of this gene does not result in reduced levels of root chemical signaling, but rather in a change in the composition of chemicals exuded from the root. Identification of this gene could allow marker-assisted breeding to generate sorghum varieties with reduced Striga infestation. Proc. Natl. Acad. Sci. U.S.A. 114 , 4471 (2017).

Seed dormancy and germination in different populations of the Argentinan endemic halophyte grass, Sporobolus phleoides (Poaceae: Chloridoideae)

Sporobolus phleoides Hack. is an endemic grass of Argentina that is considered an important genetic resource for saline environments. Knowledge of its germination behaviour is an indispensable requirement for the future potential use of this species. Thus, the effects of different factors on germination were evaluated in six representative populations collected from plants cultivated in a uniform environment. In addition, we investigated how the different parts of the seed contributed to dormancy and intraspecific variability. S. phleoides has non-deep physiological dormancy that appeared to be related to its testa. Thereby, dormancy was completely removed with puncturing treatments. High levels of germination were also obtained in seeds stratified at 8°C over 100 days in a solution of nitrates. After-ripening treatment did not have a significant effect in breaking dormancy. Similar germination behaviour and dormancy levels were observed in the different populations, especially when they were subjected to cold stratification. Although further work is needed, cold stratification appears to be the major factor in determining the time of seedling establishment in natural environments for this species.

Removal of PAHs and pesticides from polluted soils by enhanced electrokinetic-Fenton treatment

In this study, electrokinetic-Fenton treatment was used to remediate a soil polluted with PAHs and the pesticide pyrimethanil. Recently, this treatment has emerged as an interesting alternative to conventional soil treatments due to its peculiar advantages, namely the capability of treating fine and low-permeability materials, as well as that of achieving a high yield in the removals of salt content and inorganic and organic pollutants.In a standard electrokinetic-Fenton treatment, the maximum degradation of the pollutant load achieved was 67%, due to the precipitation of the metals near the cathode chamber that reduces the electro-osmotic flow of the system and thus the efficiency of the treatment. To overcome this problem, different complexing agents and pH control in the cathode chamber were evaluated to increase the electro-osmotic flux as well as to render easier the solubilization of the metal species present in the soil. Four complexing agents (ascorbic acid, citric acid, oxalic acid and ethylenediaminetetraacetic acid) in the Fenton-like treatment were evaluated. Results revealed the citric acid as the most suitable complexing agent. Thereby its efficiency was tested as pH controller by flushing it in the cathode chamber (pH 2 and 5). For the latter treatments, near total degradation was achieved after 27d.Finally, phytotoxicity tests for polluted and treated samples were carried out. The high germination levels of the soil treated under enhanced conditions concluded that nearly complete restoration was achieved.

English

The purpose of treating seeds chemically is to eradicate their pathogens and/or protect them against soil pathogens, mainly by germination time. However, there is little research on vegetables investigating the effect of this treatment on seed quality. Therefore, this study evaluates the effects of Carboxin + Thiram doses on germination and vigor of three lots of broccoli seeds, as well as on the incidence of fungi in treated seed. The 15 treatments were evaluated in a factorial system (3x5), with the first factor consisting of three lots of 'Avenger' broccoli seeds (lots 82744, 82745 and 82749), and the second factor consisting of five doses (0, 0.04, 0.06, 0.10 and 0.12% of a.i.) of Carboxin + Thiram fungicide (commercial name Vitavax-Thiran). The germination and seed vigor were evaluated, in addition to the presence of pathogens in seeds after treatment (blotter test). All lots showed high levels of germination and vigor. The lot 82749, however, showed higher value in plug test in substrate emergence (99%) than lot 82745 (95%). Regarding the treatment with Carboxin + Thiram, no changes in germination average (98%) and vigor were noticed (average for the first germination count, length, and dry weight of seedling, plug test at 10 days after sowing of 97%, 4.9 cm, 4.0 mg and 96%, respectively), showing that this fungicide, in the evaluated doses, does not affect the quality of broccoli seeds. As to seeds health, the pathogens Alternaria spp. and Fusarium spp. were detected, in addition to saprophytic species such as Penicillium, Aspergillus, Trichoderma, and Rhizopus. The higher incidence of Fusarium spp. was noticed in lot 82744, and the lowest in lot 82749. As to Penicillium spp., lot 82479 was the most contaminated. Regarding other fungi, the general incidence was very low and there was no difference between lots and doses used.   Key words: Brassica oleracea, seed treatment, fungicides.

Micro-organismos associados a sementes de pinhão-manso (Jatropha curcas L.)

Jatropha curcas is a perennial crop that stands out as a promising species for biofuel production because their grains have the potential to generate around 50% of oil. Like this culture is still little explored scientifically, the knowledge of aspects of the association of microorganisms with seeds is still incipient. The aim of this research was to evaluate the sanity and identify microorganisms associated with five ecotypes of Jatropha seeds. The treatments were different ecotypes of Jatropha seeds from different regions of the state of Minas Gerais (ecotype Uberlandia 1, ecotype Uberlandia 2, ecotype Janaúba 1, ecotype Janaúba 2 and ecotype Janaúba 3) with and without disinfection. For the evaluation of seeds were carried blotter test, test germination and seedling. It can be observed that the seeds of Jatropha analyzed showed high frequency of fungal species independent of surface disinfection. The five ecotypes of Jatropha curcas from different regions of the state of Minas Gerais showed high levels of germination, which showed no disease transmission to seedlings from the germination study.

The effect of storage temperature and duration on northern red oak acorn viability and vigour

Three separate collections of Ontario sources of northern red oak (Quercus rubra L.) acorns were made to determine the effects of long-term cold storage at +2°C, −1°C, and −2°C on their viability and vigour. We measured acorn moisture content, percent germination during storage, speed of germination and total germination values, root regrowth of seeds that germinated in storage, incidence of fungal contamination, and condition of ungerminated acorns. Viability and vigour peaked six to 12 months after acorns were placed in storage, but decreased with continued storage. After 18 months in storage, ≥60% of the acorns germinated in four of the five seedlots tested and, after 30 months in storage, ≥53% of the acorns germinated in three of the five seedlots tested. Acorn viability was only minimally affected by storage temperature; however, since temperatures above −2°C allowed acorns to germinate during storage, the preferred long-term storage temperature was −2°C. Our results suggest that, assuming proper storage conditions, most red oak seedlots will maintain relatively high germination levels when stored at −2°C for 18 months. Seedlots with particularly high initial germination and vigour may be successfully stored for up to 30 months.

Effects of seed storage on germination of two succulent desert halophytes with little dormancy and transient seed bank

Seeds of both Salsola imbricata and Haloxylon salicornicum have high germination level and germination speed, and form a transient seed bank in nature. The impacts of storage period and condition on germination level and speed were assessed in the two species. Storage for three months significantly increased both germination level and speed of seeds stored under the different conditions, compared to that of fresh seeds. In both species, nine months storage did not affect germination percentage in cold storage seeds, but completely inhibited it in field seeds. Storage for longer time in room and warm temperatures resulted in significant reduction or complete inhibition in the germination of the two species, so this was more pronounced in H. salicornicum. Storage significantly increased germinate rate index of seeds stored in all conditions till 17months in S. imbricata and till 12months in H. salicornicum. In both species, fridge storage had little effects on final germination and germination speed of seeds incubated at the different temperatures, compared to fresh seeds. However, room temperature and warm storages significantly reduced final germination and germination speed at the different temperatures, so the reduction was more pronounced at 35°C, especially in H. salicornicum.

Ozone exposure of a weed community produces adaptive changes in seed populations of Spergula arvensis.

Tropospheric ozone is one of the major drivers of global change. This stress factor alters plant growth and development. Ozone could act as a selection pressure on species communities composition, but also on population genetic background, thus affecting life history traits. Our objective was to evaluate the consequences of prolonged ozone exposure of a weed community on phenotypic traits of Spergula arvensis linked to persistence. Specifically, we predicted that the selection pressure exerted by high ozone concentrations as well as the concomitant changes in the weed community would drive population adaptive changes which will be reflected on seed germination, dormancy and longevity. In order to test seed viability and dormancy level, we conducted germination experiments for which we used seeds produced by S. arvensis plants grown within a weed community exposed to three ozone treatments during four years (0, 90 and 120 ppb). We also performed a soil seed bank experiment to test seed longevity with seeds coming from both the four-year ozone exposure experiment and from a short-term treatment conducted at ambient and added ozone concentrations. We found that prolonged ozone exposure produced changes in seed germination, dormancy and longevity, resulting in three S. arvensis populations. Seeds from the 90 ppb ozone selection treatment had the highest level of germination when stored at 75% RH and 25 °C and then scarified. These seeds showed the lowest dormancy level when being subjected to 5 ºC/5% RH and 25 ºC/75% followed by 5% RH storage conditions. Furthermore, ozone exposure increased seed persistence in the soil through a maternal effect. Given that tropospheric ozone is an important pollutant in rural areas, changes in seed traits due to ozone exposure could increase weed persistence in fields, thus affecting weed-crop interactions, which could ultimately reduce crop production.

Histidine acts as a co-germinant with glycine and taurocholate forClostridium difficilespores

It is well established that the bile salt sodium taurocholate acts as a germinant for Clostridium difficile spores and the amino acid glycine acts as a co-germinant. The aim of this study was to determine whether any other amino acids act as co-germinants. Clostridium difficile spore suspensions were exposed to different germinant solutions comprising taurocholate, glycine and an additional amino acid for 1 h before heating shocking (to kill germinating cells) or chilling on ice. Samples were then re-germinated and cultured to recover remaining viable cells. Only five amino acids out of the 19 common amino acids tested (valine, aspartic acid, arginine, histidine and serine) demonstrated co-germination activity with taurocholate and glycine. Of these, only histidine produced high levels of germination (97·9-99·9%) consistently in four strains of Cl. difficile spores. Some variation in the level of germination produced was observed between different PCR ribotypes, and the optimum concentration of amino acids with taurocholate for the germination of Cl. difficile NCTC 11204 spores was 10-100 mmol l-1. Histidine was found to be a co-germinant for Cl. difficile spores when combined with glycine and taurocholate.