Mixture toxicity of copper and zinc to barley at low level effects can be described by the Biotic Ligand Model

Background and aims

The biotic ligand model (BLM) is a bioavailability model for metals based on the concept that toxicity depends on the concentration of metal bound to a biological binding site; the biotic ligand. Here, we evaluated the BLM to interpret and explain mixture toxicity of metals (Cu and Zn).

Methods

The mixture toxicity of Cu and Zn to barley (Hordeum vulgare L.) was tested with a 4 days root elongation test in resin buffered nutrient solutions. Toxicity of one toxicant was tested in presence or absence of a low effect level of the other toxicant or in a ray design with constant toxicant ratios. All treatments ran at three different Ca concentrations (0.3, 2.2 and 10?mM) to reveal ion interaction effects.

Results

The 50 % effect level (EC50) of one metal, expressed as the free ion in solution, significantly (p?<?0.05) increased by adding a low level effect of the other metal at low Ca. Such antagonistic interactions were smaller or became insignificant at higher Ca levels. The Cu EC10 was unaffected by Zn whereas the Zn EC10 increased by Cu at low Ca. These effects obeyed the BLM combined with the independent action model for toxicants.

Conclusions

The BLM model explains the observed interactions by accounting for competition between both metals free ions and Ca²⁺ at the Cu and Zn biotic ligands. The implications of these findings for Cu/Zn interactions in soil are discussed.     

Effects of soil acidity and water stress on corn and soybean performance under a no-till system

Background and Aims

Field studies have demonstrated that aluminum (Al) toxicity is low in no-till systems during cropping seasons that have adequate and well-distributed rainfall. This study evaluated the performance of corn (Zea mays L.) and soybean (Glycine max L. Merrill) on an acid loamy soil under a long-term no-till system, in response to surface liming and as affected by genotypic tolerance to Al and water stress.

Methods

A field trial examined the effect of surface application of lime (0, 4, 8, and 12 Mg ha^?1) on no-till corn and soybean nutrition and yield. Trials were also carried out in undisturbed soil columns taken from the unlimed and limed plots. Two hybrids/cultivars of corn and soybean, one sensitive and the other moderately sensitive to Al were grown at two soil moisture levels with and without water stress (50 % and 80 % water filled pore space).

Results

Alleviating soil acidity by liming improved nutrition and increased grain yields of corn and soybean. The benefits of liming on root length density, nutrient uptake and shoot biomass production of corn and soybean were more pronounced in Al-sensitive genotypes under water stress.

Conclusions

The results suggest that plants exposed to drought stress under no-till systems are more affected by Al toxicity.     

Maintaining elevated Fe2+ concentration in solution culture for the development of a rapid and repeatable screening technique for iron toxicity tolerance in rice (Oryza sativa L.)

Background and aims

Iron toxicity decreases rice (Oryza sativa) grain yield especially in acid soils after flooding. Our aim was to establish a high-throughput screening technique using nutrient solution culture for identifying Fe-toxicity-tolerant genotypes.

Methods

Varying levels of Fe, pH, and chelators in Yoshida nutrient solution culture were tested to maintain sufficient Fe²⁺ concentration over time to optimize the severity of Fe toxicity stress for distinguishing between a tolerant (Azucena) and sensitive (IR64) genotype. The optimized solution was tested on 20 diverse genotypes in the greenhouse, with measurement of leaf bronzing scores and plant growth characteristics at the seedling stage. The same 20 genotypes were grown to maturity in a field with natural Fe toxicity stress, with measurement of seedling-stage leaf bronzing scores and grain yield to determine their inter-relationship.

Results

Optimized nutrient solution conditions were 300 mg L^?1 Fe supplied as Fe²⁺ at pH 4.0 with a 1:2 molar ratio of Fe:EDTA, which maintained sufficient Fe²⁺ stress over 5 days. The highest correlation of nutrient solution phenotypic data with field grain yield was found with leaf bronzing scores at 4 weeks, with a Pearson r of 0.628 for simple association and a Spearman corrected r of 0.610 for rank association (P?<?0.01) using 20 diverse rice genotypes with proven Fe toxicity tolerance reaction. The Leaf bronzing scores at 4 weeks in nutrient culture solution were also found highly correlated with LBS under natural field stress after 8 weeks that had highest correlation with grain yield under stress.

Conclusion

This culture solution-based standardized screening technique can be used in plant breeding programs as a high-throughput technique to identify genotypes tolerant to Fe toxicity.     

Mapping resistance genes for Oculimacula acuformis in Aegilops longissima

Key message

This study identified three QTL conferring resistance to Oculimacula acuformis in Aegilops longissima and their associated markers, which can be useful in marker-assisted selection breeding for eyespot resistance.

Abstract

Oculimacula acuformis is one of two species of soilborne fungi that cause eyespot of wheat, the other being Oculimacula yallundae. Both pathogens can coexist in the same field and produce elliptical lesions on stem bases of wheat that are indistinguishable. Pch1 and Pch2 are the only two eyespot resistance genes readily available to wheat breeders, but neither provides complete control. A new source of eyespot resistance was identified from Aegilops longissima (2n = 14, S^lS^l), a wild relative of wheat. Three QTL for resistance to O. acuformis were mapped in chromosomes 1S^l, 3S^l, and 5S^l using a recombinant inbred line population developed from the cross Ae. longissima accessions PI 542196 (R) × PI 330486 (S). The three QTL explained 66 % of phenotypic variation by β-glucuronidase score (GUS) and 84 % by visual rating. These QTL had LOD values of 10.6, 8.8, and 6.0 for GUS score, and 16.0, 10.0, and 13.0 for visual rating. QTL associated with resistance to O. acuformis have similar chromosomal locations as some for resistance to O. yallundae, except that a QTL for resistance to O. yallundae was found in chromosome 7S^l but not for O. acuformis. Thus, it appears that some genes at the same locus in Ae. longissima may control resistance to both eyespot pathogens. QTL effective against both pathogens will be most useful for breeding programs and have potential to improve the effectiveness and genetic diversity of eyespot resistance.     

Response of Populus tremula to heterogeneous B distributions in soil

Background

Poplars accumulate inordinate amounts of B in their leaves and are candidate plants for the remediation of B contaminated soil. We aimed to determine the effect of heterogeneous B distribution in soil by comparing the growth and B accumulation of young Populus tremula trees growing in soil with heterogeneous and homogeneous B distributions.

Methods

The first of two experiments focused on the tolerance and B accumulation of P. tremula under heterogeneous soil B distributions, while the second was designed to study fine root growth under such conditions in detail.

Results

Growth and B accumulation of P. tremula were unaffected by the spatial distribution of B. Root and shoot growth were both reduced simultaneously when leaf B concentrations increased above 800 mg kg^?1. In the heterogeneous soil B treatments, root growth was more reduced in spiked soil portions with B concentrations >20 mg kg^?1. Fine root length growth was stronger inhibited by B stress than secondary growth.

Conclusions

The root growth responses of P. tremula to B are primarily a systemic effect induced by shoot B toxicity and local toxicity effects on roots become dominant only at rather high soil B concentrations. Local heterogeneity in soil B should have little influence on the phytoremediation of contaminated sites.     

Effect of selenium fertilization on the accumulation of cadmium and lead in rice plants

Background and Aims

The accumulation of cadmium and lead in rice (Oryza sativa L.) grains is a potential threat to human health. In this study, the effect of selenium fertilization on the uptake and translocation of cadmium and lead in rice plants was investigated.

Methods

Rice plants were cultivated using cadmium and lead contaminated soils with selenium addition at three concentrations (0, 0.5 and 1 mg kg^?1). At maturity, plants were harvested, and element concentrations in rice tissues were analyzed by using ICP-MS.

Results

Selenium application significantly increased selenium accumulation in rice grain, and markedly decreased cadmium and lead concentrations in rice tissues. In brown rice grains, selenium application reduced cadmium concentrations by 44.4 %, but had no significant effect on lead accumulation. Selenium application significantly decreased metal mobility in soils, at 0.5 mg kg^?1 treatment, the translocation factor of cadmium and lead from soil to iron plaque decreased by 71 and 33 % respectively.

Conclusions

The mechanism of selenium mitigating of heavy metal accumulation in rice could be decreasing metal bioavailability in soil. Selenium fertilization could be an effective and feasible method to enrich selenium and reduce cadmium levels in brown rice.     

Chemical soil factors influencing plant assemblages along copper-cobalt gradients: implications for conservation and restoration

Aims

Define the chemical factors structuring plant communities of three copper-cobalt outcrops (Tenke-Fungurume, Katangan Copperbelt, D. R. Congo) presenting extreme gradients.

Methods

To discriminate plant communities, 172 vegetation records of all species percentage cover were classified based on NMDS and the Calinski criterion. Soil samples were analyzed for 13 chemical factors and means compared among communities with ANOVA. Partial canonical correspondence analysis (pCCA) was used to determine amount of variation explained individually by each factor and site effect.

Results

Seven communities were identified. Six of the studied communities were related to distinct sites. Site effect (6.0 % of global inertia) was identified as the most important factor related to plant communities’ variation followed by Cu (5.5 %), pH (3.6 %) and Co (3.5 %). Unique contribution of site effect (3.8 %) was higher than that of Cu (1.1 %) and Co (1.0 %).

Conclusions

In restoration, not only Cu and Co contents will be important to maintain vegetation diversity, attention should also be given to co-varying factors potentially limiting toxicity of metals: pH, organic matter, Ca and Mn. Physical parameters were also identified as important in the creation of adequate conditions for diverse communities. Further studies should focus on the effect of physical parameters and geology.     

Oxidation and reduction of sulfite contribute to susceptibility and detoxification of SO2 in Populus × canescens leaves

Key Message

The critical level for SO ₂ susceptibility of Populus × canescens is approximately 1.2 μL L ^?1 SO ₂ . Both sulfite oxidation and sulfite reduction and assimilation contribute to SO ₂ detoxification.

Abstract

In the present study, uptake, susceptibility and metabolism of SO₂ were analyzed in the deciduous tree species poplar (Populus × canescens). A particular focus was on the significance of sulfite oxidase (SO) for sulfite detoxification, as SO has been characterized as a safety valve for SO₂ detoxification in herbaceous plants. For this purpose, poplar plants were exposed to different levels of SO₂ (0.65, 0.8, 1.0, 1.2 μL L^?1) and were characterized by visible injuries and at the physiological level. Gas exchange parameters (stomatal conductance for water vapor, CO₂ assimilation, SO₂ uptake) of the shoots were compared with metabolite levels (sulfate, thiols) and enzyme activities [SO, adenosine 5′-phosphosulfate reductase (APR)] in expanding leaves (80–90 % expanded). The critical dosage of SO₂ that confers injury to the leaves was 1.2 μL L^?1 SO₂. The observed increase in sulfur containing compounds (sulfate and thiols) in the expanding leaves strongly correlated with total SO₂ uptake of the plant shoot, whereas SO₂ uptake rate was strongly correlated with stomatal conductance for water vapor. Furthermore, exposure to high concentration of SO₂ revealed channeling of sulfite through assimilatory sulfate reduction that contributes in addition to SO-mediated sulfite oxidation to sulfite detoxification in expanding leaves of this woody plant species.     

Molecular cloning and expression of a glycosyltransferase from <Emphasis Type="Italic">Bacillus subtilis</Emphasis> for modification of morin and related polyphenols

Objectives

To characterize glycosyltransferases from Bacillus subtilis ATCC 6633 and investigate their substrate specificity towards plant polyphenols.

Results

Among the cloned and expressed six UDP-glycosyltransferases (BsGT1-6), BsGT-1 showed activity with a wide range of polyphenols: morin, quercetin, alizarin, rehin, curcumin and aloe emodin. The gene of BsGT-1 has an ORF of 1206 bp encoding 402 amino acids. The recombinant enzyme was purified to homogeneity by Ni–NTA affinity chromatograph, and its biochemical characteristics were identified by HPLC–UV/MS, ¹H-NMR and ¹³C-NMR. BsGT-1 has an MW of approx. 46 kDa as indicated by SDS-PAGE; its activity was optimal at 40 °C and pH 8.5. The Km value of BsGT-1 towards morin was 110 μM.

Conclusions

BsGT-1 from B. subtilis was cloned. It had high catalytic capabilities towards polyphenols which would make it feasible for the structural modification of polyphenols.

     

Reducing basal salicylic acid enhances Arabidopsis tolerance to lead or cadmium

Aims

Phytoremediation is an emerging strategy for the removal of heavy metal contaminants. However, one of the prerequisite is to understand adequately plant resistant mechanisms. The present study was performed to assess the role of endogenous SA in plant response to Pb or Cd using wild-type (wt) Arabidopsis and its SA-accumulating mutant snc1, SA-reducing transgenic line nahG, SA signal-blocking npr1-1, and snc1/nahG (i.e. expression of nahG in snc1 plant) with a comparable level of SA to the wt.

Methods

Plants were grown hydroponically in controlled conditions. For heavy metal exposure, Pb²⁺ or Cd²⁺ at final concentrations of 50 μM, 100 μM, and 150 μM, respectively, was added to the culture solution. Unless otherwise indicated, samples were harvested after 7 d of exposure, and used for analyses.

Results

Compared to the wt level, the high endogenous SA significantly potentiated Pb- and Cd-induced plant growth retardation, whereas SA deficiency decreased the growth inhibition, and SA signaling blockage also had some protective effect. The expression of nahG in snc1 plant mitigated effectively the growth inhibition. The SA-related mechanism was involved in redox homeostasis, photosynthetic process, and soluble matter accumulation.

Conclusions

These results suggest that Pb- or Cd-induced phytotoxicity in Arabidopsis was intensified by elevated endogenous SA, whereas ameliorated by reduced SA.     

Overexpression of citrate operon in Herbaspirillum seropedicae Z67 enhances organic acid secretion,mineral phosphate solubilization and growth promotion of Oryza sativa

Background and aims

Herbaspirillum seropedicae Z67, nitrogen fixing endophyte, significantly promotes the growth of cereals. Organic acid secreting nitrogen fixing rhizobacteria have better plant growth promotion abilities due to mineral phosphate solubilization.

Method

Plasmids pAB7, pJNK3 and pJNK4 containing Escherichia coli cs (gltA), NADH insensitive cs (gltA1), and citrate operon consisting of gltA1 gene along with Salmonella typhimurium Na⁺ dependent citrate transporter (citC) gene under constitutive lac promoter were constructed in broad host range plasmid pUCPM18-Km^r. The plasmid transformants of H. seropedicae Z67 were obtained by electroporation.

Results

Hs (pAB7) and Hs (pJNK3) had increased CS activity but citric acid secretion was not significant. Hs (pJNK3) secreted 45 mM acetic acid while Hs (pJNK4) secreted 2.7 mM citric and 51 mM acetic acids. Hs (pJNK3) and Hs (pJNK4) released 80 μM and 110 μM amount of P from rock phosphate, respectively, in buffered medium under both aerobic and micro aerobic conditions. These transformants showed better plant growth promoting factors. Upon inoculation to rice plants (Gujarat – 17), increase of Fresh weight, Dry weight N, P and K content was observed.

Conclusion

Thus the study demonstrates that artificial citrate operon in H. seropedicae Z67 enhances phosphate solubilization and plant growth promotion abilities.     

Assessment of nitrate leaching loss on a yield-scaled basis from maize and wheat cropping systems

Background and aims

It is so far a gap in knowledge to assess nitrate (NO₃ ^?) leaching loss linking with crop yield for a given cereal cropping system.

Methods

We conducted a meta-analysis on 32 published studies reporting both NO₃ ^? leaching losses and crop yields in the maize (N?=?20) and wheat (N?=?12) systems.

Results

On average, 22 % and 15 % of applied fertilizer N to wheat and maize systems worldwide are leached in the form of NO₃ ^?, respectively. The average area-scaled NO₃ ^- leaching loss for maize (57.4 kg N ha^?1) was approx. two times higher than for wheat (29.0 kg N ha^?1). While, if scaled to crop yields, the average yield-scaled NO₃ ^? losses were comparable between maize (5.40 kg N Mg^?1) and wheat (5.41 kg N Mg^?1) systems. Across all sites, the lowest yield-scaled NO₃ ^? leaching losses were observed at slightly suboptimal fertilization rates, corresponding to 90 % and 96 % of maximum maize or wheat yields, respectively.

Conclusions

Our findings suggest that small adjustments of agricultural N management practices can effectively reduce yield-scaled NO₃ ^? leaching losses. However, further targeted field experiments are still needed to identify at regional scale best agricultural management practices for reducing yield-scaled NO₃ ^? leaching losses in maize and wheat systems.     

Time change of aluminium toxicity in the acid bulk soil and the rhizosphere in Norway spruce (Picea abies (L.) Karst.) and beech (Fagus sylvatica L.) stands

Context

In acidic forest soils, aluminium can alter tree health due to its potential toxicity. Aluminium phytotoxicity is mainly influenced by its chemical form and its availability.

Methods

As physical-chemical indicators of Al toxicity in soil, Al speciation in soil solutions and in the exchange complex was measured in the rhizosphere and the bulk soil of two tree species (Norway spruce (Picea abies (L.) Karst.) and European Beech (Fagus sylvatica L.) in an acidic soil and in 4 months (November, February, May and August) representing the four seasons in a year.

Results

In the bulk soil, Al toxicity was generally higher under Norway spruce than under beech. Furthermore, temporal changes in Al behaviour were identified under Norway spruce but not under beech. The monomeric Al in the soil solutions and the exchangeable Al in the solid soil increased significantly in February under Norway spruce and were positively correlated with nitrate concentration, suggesting that nitrate influence Al speciation and mobility under Norway spruce. In the rhizosphere, Al toxicity was restricted through Al complexation by organic compounds and by nutrient contents independently from the season. The ecological importance of the rhizosphere in Al detoxification is discussed.

Conclusions

This study suggests that plant specific differences as well as seasonal changes in plant physiology, microbial activity and microclimatology influence aluminum toxicity in acid forest soils.     

Stable carbon and nitrogen isotope ratios of Eucalyptus and Acacia species along a seasonal rainfall gradient in Western Australia

Key message

Eucalyptus and Acacia species were surprisingly similar with respect to variations in δ ¹³ C, δ ¹⁵ N. Both genera respond with speciation and associated changes in leaf structure to drought.

Abstract

Stable carbon and nitrogen isotope ratios (δ¹³C and δ¹⁵N) in leaves of eucalypts (Corymbia and Eucalyptus) and Acacia (and some additional Fabaceae) species were investigated together with specific leaf area (SLA), leaf nitrogen (N) and leaf phosphorous (P) concentration along a north–south transect through Western Australia covering winter- and summer-dominated rainfall between 100 and 1,200 mm annually. We investigated 62 eucalypts and 78 woody Fabaceae species, mainly of the genus Acacia. Leaf δ¹³C values of Eucalyptus and Acacia species generally increased linearly with latitude from ?29.5 ± 1.3 ‰ in the summer-dominated rainfall zone (15°S–18°S) to about ?25.7 ± 1.1 ‰ in the winter-dominated rainfall zone (29°S–31°S). δ¹⁵N increased initially with southern latitudes (0.5 ± 1.6 ‰ at 15°S; 5.8 ± 3.3 ‰ at 24–29°S) but decreased again further South (4.6 ± 3.5 ‰ at 31°S). The variation in δ¹³C and δ¹⁵N was probably due to speciation of Eucalyptus and Acacia into very local populations. There were no species that were distributed over the whole sampling area. The variation in leaf traits was larger between species than within species. Average nitrogen concentrations were 11.9 ± 1.05 mg g^?1 in Eucalyptus, and were 18.7 ± 4.1 mg g^?1 in Acacia. Even though the average nitrogen concentration was higher in Acacia than Eucalyptus, δ¹⁵N gave no clear indication for N₂ fixation in Acacia. In a multiple regression, latitude (as a surrogate for rainfall seasonality), mean rainfall, leaf nitrogen concentration, specific leaf area and nitrogen fixation were significant and explained 69 % of the variation of δ¹³C, but only 36 % of the variation of δ¹⁵N. Higher nitrogen and phosphorus concentration could give Acacia an advantage over Eucalyptus in arid regions of undefined rainfall seasonality.     

Characterization of a recombinant l-fucose isomerase from Caldicellulosiruptor saccharolyticus that isomerizes l-fucose, d-arabinose, d-altrose, and l-galactose

A recombinant l-fucose isomerase from Caldicellulosiruptor saccharolyticus was purified as a single 68 kDa band with an activity of 76 U mg^?1. The molecular mass of the native enzyme was 204 kDa as a trimer. The maximum activity for l-fucose isomerization was at pH 7 and 75°C in the presence of 1 mM Mn²⁺. Its half-life at 70°C was 6.1 h. For aldose substrates, the enzyme displayed activity in decreasing order for l-fucose, with a k _cat of 11,910 min^?1 and a K _m of 140 mM, d-arabinose, d-altrose, and l-galactose. These aldoses were converted to the ketoses l-fuculose, d-ribulose, d-psicose, and l-tagatose, respectively, with 24, 24, 85, 55% conversion yields after 3 h.     

Population structure and association mapping of yield contributing agronomic traits in foxtail millet

Key message

Association analyses accounting for population structure and relative kinship identified eight SSR markers ( p < 0.01) showing significant association ( R ²  = 18 %) with nine agronomic traits in foxtail millet.

Abstract

Association mapping is an efficient tool for identifying genes regulating complex traits. Although association mapping using genomic simple sequence repeat (SSR) markers has been successfully demonstrated in many agronomically important crops, very few reports are available on marker-trait association analysis in foxtail millet. In the present study, 184 foxtail millet accessions from diverse geographical locations were genotyped using 50 SSR markers representing the nine chromosomes of foxtail millet. The genetic diversity within these accessions was examined using a genetic distance-based and a general model-based clustering method. The model-based analysis using 50 SSR markers identified an underlying population structure comprising five sub-populations which corresponded well with distance-based groupings. The phenotyping of plants was carried out in the field for three consecutive years for 20 yield contributing agronomic traits. The linkage disequilibrium analysis considering population structure and relative kinship identified eight SSR markers (p < 0.01) on different chromosomes showing significant association (R ² = 18 %) with nine agronomic traits. Four of these markers were associated with multiple traits. The integration of genetic and physical map information of eight SSR markers with their functional annotation revealed strong association of two markers encoding for phospholipid acyltransferase and ubiquitin carboxyl-terminal hydrolase located on the same chromosome (5) with flag leaf width and grain yield, respectively. Our findings on association mapping is the first report on Indian foxtail millet germplasm and this could be effectively applied in foxtail millet breeding to further uncover marker-trait associations with a large number of markers.     

Significance of Arg3, Arg54, and Tyr58 of l-aspartate α-decarboxylase from Corynebacterium glutamicum in the process of self-cleavage

Purpose of work

We have elucidated the significance of three key amino acid residues of l-aspartate α-decarboxylase that act remotely from its cleavage site for its functional self-cleavage as well as for its catalytic activity. These results provide useful fundamental information for engineering l-aspartate α-decarboxylase. l-Aspartate α-decarboxylase (ADC) from Corynebacterium glutamicum, and encoded by panD, was cloned and expressed in Escherichia coli and then purified. Three amino acid residues were found to be related to ADC self-cleavage. Mutating R3 to either A, Q, N, L, D, or E produced only the unprocessed pro-enzyme. Although mutating R54 and Y58 into A or K and A or T, respectively, partly influenced ADC self-cleavage, the specific activity of each of the four ßmutants decreased to 3.5, 4, 2.4, and 2.6 U mg^?1, respectively, compared with a specific activity of 690 U mg^?1 for the wild-type enzyme. Thus, R3 triggers ADC self-cleavage and completes the modification of the active site with assistance by R54 and Y58. These results will help to engineer ADC for improved industrial applications.     

Rhizobial counts in peat inoculants vary amongst legume inoculant groups at manufacture and with storage: implications for quality standards

Background and aims

Inoculation of legumes at sowing with rhizobia has arguably been one of the most cost-effective practices in modern agriculture. Critical aspects of inoculant quality are rhizobial counts at manufacture/registration and shelf (product) life.

Methods

In order to re-evaluate the Australian standards for peat-based inoculants, we assessed numbers of rhizobia (rhizobial counts) and presence of contaminants in 1,234 individual packets of peat–based inoculants from 13 different inoculant groups that were either freshly manufactured or had been stored at 4 °C for up to 38 months to determine (a) rates of decline of rhizobial populations, and (b) effects of presence of contaminants on rhizobial populations. We also assessed effects of inoculant age on survival of the rhizobia during and immediately after inoculation of polyethylene beads.

Results

Rhizobial populations in the peat inoculants at manufacture and decline rates varied substantially amongst the 13 inoculant groups. The most stable were Sinorhizobium, Bradyrhizobium and Mesorhizobium with Rhizobium, particularly R. leguminosarum bv. trifolii the least stable. The presence of contaminants at the 10^?6 level of dilution, i.e. >log 6.7 g^?1 peat, reduced rhizobial numbers in the stored inoculants by an average of 37 %. Survival on beads following inoculation improved 2–3 fold with increasing age of inoculant.

Conclusions

We concluded that the Australian standards for peat-based rhizobial inoculants should be reassessed to account for the large differences amongst the groups in counts at manufacture and survival rates during storage. Key recommendations are to increase expiry counts from log 8.0 to log 8.7 rhizobia g^?1 peat and to have four levels of inoculant shelf life ranging from 12 months to 3 years.