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Growth and development of Lotus and Trifolium species under saline and waterlogging conditions

Angus Galloway1, Joshua Cables1, David Parsons1,2, Peter Lane1,2 and Eric Hall2

1 School of Agricultural Science, The University of Tasmania, Private Bag 54, Hobart TAS 7001,
Tasmanian Institute of Agricultural Research, The University of Tasmania, Private Bag 54, Hobart TAS 7001,


Saline and waterlogging conditions can be detrimental to the growth of pasture plants. The aim of this study was to compare the tolerance of two new pasture species, Lotus tenuis and Trifolium tumens, to comparable traditional species, L. corniculatus, L. pedunculatus, T. repens, and T. fragiferum. Plant growth was studied under four treatments; control (non-saline, non-waterlogging), saline (150mM), waterlogging, and saline with waterlogging. The waterlogging treatment was imposed by placing the pots in plastic troughs where the water level was maintained at 2 cm below the rim. Root and shoot dry matter yield, and root length were measured 27 days after emergence. There was no interaction (P<0.05) between salinity and waterlogging. Waterlogging had a negative effect on root length for all species, but not on shoot or root dry matter. Of the Lotus species, the four accessions of L. tenuis were not affected by salinity, whereas salinity reduced shoot dry matter yield in the accessions of L. corniculatus, L. pedunculatus and both Trifolium species. Although there was a 31% reduction in L. corniculatus shoot dry matter yield with the salinity treatment, it was the highest yielding species for this treatment. Salinity reduced root dry matter yield in T. tumens and T. repens, further confirming their sensitivity to salinity. The results confirm that L. tenuis has tolerance of saline and waterlogging conditions where it has potential to be a useful species.

Key Words

Trifolium tumens, Lotus tenuis, Lotus corniculatus, Lotus pedunculatus


Salinity is an ever increasing problem in Australia, with over 2 million hectares of agricultural land reported as being affected by salinity (Australian Bureau of Statistics 2002). In addition to this problem, plants can be exposed to waterlogging during certain times of the year. The combined effect of salinity and waterlogging can lead to extremely poor productivity from pasture legumes. Traditional clover varieties such as Trifolium fragiferum (strawberry clover) and Trifolium repens (white clover) are resistant to waterlogging (Gibberd et al. 2001). T. fragiferum shows moderate salinity tolerance (Rumbaugh et al. 1993), whereas T. repens is susceptible to salinity (Rogers et al. 1997).

There are several published papers pertaining to the waterlogging tolerance of the three species of Lotus that were grown in this study, (Gibberd et al. 2001; Striker et al. 2005) but there has been less research on the combined effects of salinity and waterlogging (Teakle et al. 2006). As a newly domesticated species, there has also been very little work done on Trifolium tumens, and no indication in the literature of its salinity or waterlogging tolerance. The objective of this project was to determine if there is an interaction between salinity and waterlogging for a range of clover and lotus species.


Seed was supplied by the TIAR genetic resources collection and Tasglobal Seeds and consisted of three Lotus and three Trifolium species. The Lotus species included L. corniculatus (Tas 2948), L. pedunculatus (Tas 280), and L. tenuis (Tas 2527, Tas 2828, Tas 2849, Tas 2526). The Trifolium species were T. repens (Tas 2861), T. fragiferum (Palestine) and a new cultivar of T. tumens (Permatas). The seeds were all scarified to counter hard seededness, then placed on filter paper in petri dishes with distilled water and germinated in a 21oC room for three days. Six germinated seeds were planted into 100mm pots containing a 30 percent perlite and 70 percent sterilized sand mix along with 5 ml of Osmocote slow release fertiliser (Scotts Australia 2009) on top of the growing medium. Plants were grown under overhead irrigation for 28 days and a full strength Hoagland nutrient solution was applied to all pots three times per week.

The experiment was set up in a randomized complete block design with nine species/accessions, four salinity and waterlogging treatments and four replicates. Each experimental unit consisted of one pot with four seedlings of a single species/accession. The treatments included salinity at 150 mM, waterlogging, 150 mM salinity and waterlogging combined, and a control. The waterlogging treatment was imposed for the duration of the trial by placing the pots in plastic troughs. A gravity-fed irrigation system with an overflow pipe in each trough was used to maintain the water level at approximately 2 cm below the rim. All troughs were drained and replenished halfway through the trial to eliminate the potential of nutrients, especially salts, concentrating in the water supply. After 27 days of imposing the treatments the plants were harvested and analyzed for shoot and root dry matter (DM) yield, and root length. All experiments were analysed using PROC GLM in SAS, v. 9.1 (SAS Institute, 2003) and Fisher’s LSD was used to test the differences (P≤0.05) among means.


There was a significant effect of waterlogging on the root length, but not on shoot or root weight (Table 1). The mean root length decreased from 176 mm in the control treatment to 158 mm in the waterlogged treatment.

Table 1. The significance of treatments on shoot DM yield, root DM yield and the longest root length.


Shoot DM yield (g)

Root DM yield (g)

Root Length (mm)

Salinity (S)




Waterlogging (W)




Accession (A)




S x W




S x A




W x A




S x W x A




NS, Not significant; * P <0.05; ** P<0.01; *** P<0.001.

For all measured variables there was a significant interaction between salinity and accession (Table 1). The interactive effect of salinity and accession on shoot DM yield is shown in Figure 1. For shoot DM yield of L. tenuis accessions there was no significant difference between the control and salinity treatments and they were the lowest yielding accessions under the control treatment. L. corniculatus had the third highest shoot DM yield under the control treatment. For all species except L. tenuis there was a significant decline in shoot DM yield under saline conditions. The shoot DM yield of L. pedunculatus under saline conditions was about half that of the control. Shoot DM yield of all the Trifolium cultivars was greatly affected by the salinity treatments imposed, particularly T. tumens which experienced a decrease of 75%.

Figure 1. The interaction of salinity and cultivar on shoot dry matter after 27 days of imposed stress. Indicated significance levels are the comparison between the control and salinity treatment for each cultivar (NS, Not significant; * P <0.05; ** P<0.01; *** P<0.001)

Figure 2. The interaction of salinity and cultivar on root dry matter after 27 days of imposed stress. Indicated significance levels are the comparison between the control and salinity treatment for each cultivar (NS, Not significant; * P <0.05; ** P<0.01; *** P<0.001)

The interaction of salinity and cultivar on root DM yield is shown in Figure 2. The results were similar to shoot DM yield with no effect of salinity on root DM yield for L. tenuis accessions, L. corniculatus, L. pedunculatus or T. fragiferum. Root DM yield under the salinity treatment was reduced in T. repens by approximately 50% and in T. tumens by approximately 70%.

There was no significant effect of salinity on root length for L. pedunculatus, L. tenuis or T. fragiferum; whereas the root length of L. corniculatus, T. repens and T. tumens was slightly reduced by the salinity treatment (Figure 3).

Figure 3. The interaction of salinity and cultivar on the longest root length after 27 days of imposed stress. Indicated significance levels are the comparison between the control and salinity treatment for each cultivar

(NS, Not significant; * P <0.05; ** P<0.01; *** P<0.001)

Discussion and Conclusion

The results confirm that the effect of salinity differs between species. L. tenuis was least affected by salinity with no effect on shoot dry matter yield, root dry matter yield, or shoot length. This result relates well with conclusions made by Teakle et al. (2006) that showed that L. tenuis is highly salt tolerant. L. corniculatus was less salt tolerant than L. tenuis but more tolerant than L. pedunculatus. The commercially available Trifolium species performed as described in previous research under saline conditions (Rumbaugh et al. 1993; Rogers et al. 1997). The growth in plants of T. repens were significantly affected by salinity, while only the shoot dry matter yield of T. fragiferum was affected. For T. tumens, shoot DM yield, root DM yield, and root length were all significantly affected by salinity, and thus it can be concluded that the species is not salt tolerant.

There was no interaction between waterlogging and salinity, a result which is contrary to (Teakle et al. 2006) who documented that L. tenuis had a greater tolerance to the combined effects of waterlogging and salinity when compared to L. corniculatus.

Future research opportunities would benefit by using different levels of salinity to determine the boundaries of salt tolerance for the different species and by including, waterlogging-sensitive “check” species such as lucerne (Real et al. 2008), to provide a comparison with the experimental species. In addition, when studying perennial species such as Lotus, it may be necessary to impose the waterlogging treatment for longer periods (greater than 27 days) in order to determine genotypic differences in growth.


This experiment was performed by students of a third year agronomy unit at the University of Tasmania. We would like to thank the following people for their help and assistance in making this experiment possible. TasGlobal Seeds for supplying the seed, Phil Andrews for assistance in the university glasshouse, and Assoc. Prof. Sergey Shabala for discussions in relation to salinity concentration.


Australian Bureau of Statistics (2002). Salinity on Australian Farms. Australian Bureau of Statistics Canberra. 4615.0. e3c62b38c2b153aeca256c8b0081eb9b!OpenDocument#Map%20of%20NAP%20Regions. Accessed 25 June 2009.

Gibberd M, Gray J, Cocks P and Colmer T (2001). Waterlogging tolerance among a diverse range of Trifolium accessions is related to root porosity, lateral root formation and aerotropic rooting. Annals of Botany 88(4), 579-589.

Real D, Warden J, Sandral GA and Colmer TD (2008). Waterlogging tolerance and recovery of 10 Lotus species. Australian Journal of Experimental Agriculture 48(4), 480-487.

Rogers M, Noble C, Halloran G and Nicolas M (1997). Selecting for salt tolerance in white clover (Trifolium repens): Chloride ion exclusion and its heritability. New Phytologist 135(4), 645-654.

Rumbaugh M, Pendery B and James D (1993). Variation in the salinity and waterlogging tolerance of strawberry clover (Trifolium fragiferum L.). Plant and Soil 153(2), 265-271.

Scotts Australia (2009). "Osmocote Total All Purpose." Retrieved 1/06, 2009, from

Striker GG, Insausti P, Grimoldi A, Ploschuk E and Vasellati V (2005). "Physiological and anatomical basis of differential tolerance to soil flooding of Lotus corniculatus L. and Lotus glaber mill. Plant and Soil 276(1-2), 301-311.

Teakle NL, Real D and Colmer T (2006). Growth and ion relations in response to combined salinity and waterlogging in the perennial forage legumes Lotus corniculatus and Lotus tenuis. Plant and Soil 289(1-2), 369-383.

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