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Botanical Composition Improvement with Subterranean Clover (Trifolium subterraneum L.) in Grass Mixtures

Viliana Vasileva
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Botanical composition of cocksfoot and tall fescue, pure and in mixtures with three subspecies subterranean clover were investigated in field trial in the Institute of Forage Crops, Pleven, Bulgaria (2011-2013). The next ratios were used: Grass+Trifolium subterraneum ssp. brachycalicinum (50:50%), Grass+Trifolium subterraneum ssp. yaninicum (50:50%), Grass+Trifolium subterraneum ssp. subterraneum (50:50%), Grass+Trifolium subterraneum ssp. brachycalicinum+Trifolium subterraneum ssp. yaninicum+Trifolium subterraneum ssp. subterraneum (25:25:25:25%). It was found that inclusion the subterranean clover as a component of mixtures with cocksfoot and tall fescue had positive effect on the botanical composition and decreased the weed infestation in the swards. Trifolium subterraneum ssp. brachycalicinum in mixture with cocksfoot had faster growing rate as well the biggest part in the sward (31.0%) and reduced the weeds half. Trifolium subterraneum ssp. brachycalicinum in mixtures with tall fescue had small part and decreased the weeds by 28.1%. Subterranean clover as a legume component contributed to increasing the leaf/stem ratio of cocksfoot and tall fescue.

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Viliana Vasileva , 2016. Botanical Composition Improvement with Subterranean Clover (Trifolium subterraneum L.) in Grass Mixtures. Journal of Applied Sciences, 16: 68-76.

DOI: 10.3923/jas.2016.68.76

Received: November 09, 2015; Accepted: December 21, 2015; Published: January 15, 2016


Interest towards intercropped systems based on co-cultivation of legumes and cereals is rising in recent years due to their role in formation of a system for sustainable and organic farming (Pypers et al., 2005; Luscher et al., 2014; Kusvuran et al., 2014). It has been known long ago that cereal/legume intercropping systems are highly efficient in relation to the using of sources of environment than separately grown crops (Porqueddu et al., 2003; Vasilev et al., 2005; Peyraud et al., 2009; Mahapatra, 2011). The leguminous component in the intercropping system contributes for the increasing of nitrogen content in the system due to its nitrogen fixing ability providing the majority of nitrogen required for them as well for the accompanying grass component (Pypers et al., 2005; Peeters et al., 2006; Pozdisek et al., 2011; Vanlauwe et al., 2014).

The permanent changes having occurred in the last ten-year periods in the climate (increase of average yearly temperatures, long droughts in the spring and summer, increase of CO2 concentration in the atmosphere) present a serious risk to the perennial crops because affect their capacity for several years. This requires to study new herbaceous forage species having pronounced resistance to unfavorable abiotic factors and good adaptive capacity towards the new conditions (Gornall et al., 2010; Bostan et al., 2013; Mihovski and Kirilov, 2014; Luscher et al., 2014; Aranjuelo et al., 2014). At present the main interest in directed towards more drought resistant and drought tolerant components. Legumes species that can provide self-sowing and persist continuously in the sward become of practical importance (Carneiro, 1999).

Subterranean clover (Trifolium subterraneum L.) is an annual drought resistant ephemeral legume with winter-spring type of development and ability for self-sowing (Yakimova and Yancheva, 1986; Piano et al., 1996; Frame et al., 1998). It is a widespread component in the pastures and other grasslands of the temperate areas of central and Northern Europe and America (Frame et al., 1998; Pecetti and Piano, 1998, 2002; Kyriazopoulos et al., 2008). The species is found In Bulgaria in open, dry grasslands in the plains and lowlands (Assyov et al., 2012).

An important element of the biology of this species is that its reproductive organs are formed in early may and the seeds ripen before the end of the spring in hedgehog-shaped heads that remain on the soil surface. Heads are 10.0-15.0 mm spherical, loose, appressed on the soil and self-burying (Kozhuharov, 1976). A big part of the seeds germinated during the autumn. Other part germinated during the spring of the next year but because of the winter-spring type of the development the plants took part in the sward during most of the vegetation season, some of them wintering and on the next spring formed seeds (Frame et al., 1998; Vasilev, 2006). Substantial part of the formed seeds is solid and germinates after two-three years. This biological specificity turns the superficial soil layer into an original seed bank which makes the species even more flexible one (Pecetti and Piano, 1994). The precipitations during the late summer contribute to emergence of new self-sown plants (Frame et al., 1998). So, subterranean clover although an annual species due to its biological ability to self-seeding present in the sward composition at the beginning and end of the vegetation. The annual seed formation and germination is a reason the crop of subterranean clover to be appear as perennial one (Vasilev, 2006).

Investigations on subterranean clover as a component of sown pasture in recent years have shown that it has practical application for local climatic conditions in Bulgaria (Vasilev, 2006, 2009; Ilieva and Vasileva, 2011; Vasileva et al., 2011; Vasileva and Vasilev, 2012; Vasileva, 2014, 2015a, b; Ilieva et al., 2015). When sown at an appropriate time in the autumn, it establishes a uniform stand before the beginning of the permanent cold spell and grows up early in the spring and forms a dense sward. A marked winter resistance, effective utilization of autumn-winter soil moisture, successful seed formation and self-sowing at the end of spring allow subterranean clover to avoid summer droughts. Subterranean clover is a suitable component for the mixture with grasses. In the present work subterranean clover was studied in mixture with cocksfoot and tall fescue.

Cocksfoot is a deeply-rooted medium to longlived highly productive grass species. It initiates growth early in the spring and due to the deep roots and access moisture in lower soil profiles, grows well during hot summer months as well during the autumn (Jacobs and Siddoway, 2007).

Tall fescue has high resistance to high summer temperatures, withstands dry as well wet soils and is a suitable component for mixtures with hay-pasture direction of use (Katova, 2007).

The aim of this work to study the botanical composition of mixtures of cocksfoot and tall fescue with subterranean clover.


The trial was conducted at the experimental field (43°23’N, 24°34’E, 230 m altitude) of the Institute of Forage Crops, Pleven, Bulgaria (2011-2013) on podzolized soil subtype without irrigation. Cocksfoot (Dactylis glomerata L.) and tall fescue (Festuca arundinacea schreb.) pure and in mixture with three subclover subspecies, i.e., Trifolium subterraneum ssp. brachycalicinum, Trifolium subterraneum ssp. yaninicum and T. subterraneum ssp. subterraneum were tested. The trial was carried out on the long plot method with size of plots 70 m2 four replicated. The variants were as follows: Cocksfoot (100%), cocksfoot+Trifolium subterraneum ssp. brachycalicinum (50:50%), cocksfoot+Trifolium subterraneum ssp. yaninicum (50:50%), cocksfoot+Trifolium subterraneum ssp. subterraneum (50:50%), cocksfoot+Trifolium subterraneum ssp. brachycalicinum+Trifolium subterraneum ssp. yaninicum+Trifolium subterraneum ssp. subterraneum (25:25:25:25%), tall fescue (100%), tall fescue+Trifolium subterraneum ssp. brachycalicinum (50:50%), tall fescue+Trifolium subterraneum ssp. yaninicum (50:50%), tall fescue+Trifolium subterraneum ssp. subterraneum (50:50%), tall fescue+Trifolium subterraneum ssp. brachycalicinum+Trifolium subterraneum ssp. yaninicum+Trifolium subterraneum ssp. subterraneum (25:25:25:25%).

Cocksfoot cv. "Dabrava" and tall fescue cv. "Albena" were used. Subclover for: Trifolium subterraneum ssp. brachycalicinum-cv. "Antas", Trifolium subterraneum ssp. yaninicum-cv. "Trikkala" and Trifolium subterraneum ssp. subterraneum-cv. "Denmark".

The sowing was done on September, 17, 2011 between row space was 11.5 cm and sowing rates: Tall fescue 25 kg ha–1, cocksfoot 25 kg ha–1, subterranean clover -25 kg ha–1. No fertilizers and pesticides were applied during the vegetation. The swards were harvested for forage. To determine botanical composition samples before the cutting was taken from the all replications using 0.25 m2 quadrate. Leaf/stem ratio of the grasses (pure and in mixtures) based on fresh weight was calculated.

In the present work the data from one cut harvested on June 6, 2012 and two cuts harvested on May 7 and July 10, 2013. Experimental data for leaf/stem ratio were statistically processed using SPSS (2012).


The period of study was characterized as an unfavorable in regard to the agro meteorological conditions. Data for mean air temperature and cumulative rainfall was shown on the Table 1. After the sowing long dry period occur. Extremely high temperatures affected the growth and development of the plants, botanical composition and productivity of the swards. One cut only was harvested.

Botanical composition of the swards is one of the main criteria for their resistance (Mannetje, 2003).

Cocksfoot and mixtures with subterranean clover: Cocksfoot has slowly growing rate during the first year and attacked by weeds (Vasilev, 2000; Kirwan et al., 2005; Jacobs and Siddoway, 2007). Regardless the autumn sowing cocksfoot it could not tiller to the frosts appearing. Plant development during the vegetation was slowly and weeds infestation significant (41.0%) (Fig. 1).

Part of the cocksfoot in mixture was affected by the growing rate of subterranean clover.

Despite the unfavorable conditions subterranean clover well developed in all swards tested. Comparing the three subspecies it is apparent that Trifolium subterraneum ssp. brachycalicinum took 31.0% in the mixture and weed infestation was half reduced as compared to the pure sward.

Table 1:Meteorological conditions for the period of study (2011-2013)

Fig. 1: Botanical composition of cocksfoot pure and in mixture with subterranean clover (one year after sowing)

Similar were results of Porqueddu et al. (2003). Significantly smaller was the part of Trifolium subterraneum ssp. yaninicum and Trifolium subterraneum ssp. subterraneum, 14.3 and 5.5%, respectively and the weeds were 32.3 and 39.0%, i.e. as the control. In the four-component mixture subterranean clovers part was 23.8 and 15.0% accounted for Trifolium subterraneum ssp. brachycalicinum. Significantly slower growth rates were found for both, Trifolium subterraneum ssp. yaninicum and Trifolium subterraneum ssp. subterraneum and they suppress the weeds less as compared to Trifolium subterraneum ssp. brachycalicinum.

Rainfall in the third year of the experiment was unevenly distributed. An early spring drought was occurred and lasts one month (from April 18 to May 16). The average daily and average monthly average maximal temperatures were above the norm. Moisture in the top soil was insufficient for the normal development of the crops as a whole, especially grasses.

Subterranean clover regardless the shallow root system as compared to perennial legumes (alfalfa, birdsfoot trefoil, sainfoin) rapidly developed in the first days of the year suitable for vegetation. Subterranean clover covered the soil surface and due to the prostrate habit retains the moisture which influences the accompanying component in the mixture.

Data for botanical composition of the swards in first cut of the second year showed that the weed infestation was considerably less (Fig. 2). Most weeds were found in the pure sward.

The effect of the subspecies of the clover in the mixture was significant. Trifolium subterraneum ssp. brachycalicinum which characterized with fast growing rate took part of 28.5%. The other two subspecies subterranean clover formed smaller biomass 15.1 and 11.3%, respectively for Trifolium subterraneum ssp. yaninicum and Trifolium subterraneum ssp. subterraneum and did not affect the weeds. The part of subterranean clover in four-component mixture was 25.4 and 12.2% of them occupied Trifolium subterraneum ssp. brachycalicinum.

Since the second half of May when the second cut is forming, rainfall was evenly distributed which favored the development of crops. Subterranean clover’s part in the second cut was up 10.7% and there were no significant differences for Trifolium subterraneum ssp. brachycalicinum and Trifolium subterraneum ssp. yaninicum. The smallest part was found for Trifolium subterraneum ssp. subterraneum which is less competitive. As a whole the part of subterranean clovers in the second cut was half reduced as compared to first cut due to the biology of the crop.

Tall fescue and mixtures with subterranean clover: Data for botanical composition of the pure sward of tall fescue during the first year after sowing showed that grass predominated 57.3% (Fig. 3).

Subterranean clover develops less in mixtures with tall fescue and its part for Trifolium subterraneum ssp. brachycalicinum reached up 11.0%. Smaller was the share for Trifolium subterraneum ssp. yaninicum (8.8%) and Trifolium subterraneum ssp. subterraneum (3.7%).

Fig. 2(a-b): Botanical composition of (a) Cocksfoot pure and in (b) mixture with subterranean clover (two years after sowing)

Fig. 3: Botanical composition of tall fescue pure and in mixture with subterranean clover (one year after sowing)

Subterranean clover has no effect on the part of tall fescue in the sward and it remains almost unchanged, varying in narrow limits (50.0-59.0%). In the mixtures with Trifolium subterraneum ssp. brachycalicinum the reduction of weeds was by 28.1% and almost 50.0% in mixtures with cocksfoot.

Fig. 4(a-b): Botanical composition of (a) Tall fescue pure and in (b) mixture with subterranean clover (second year after sowing)

Nearly, 1/4 falls on the subterranean clovers in the four-component mixture.

Tall fescue (pure) predominated (88.5%) in first cut during the second year after sowing (Fig. 4). Part of Trifolium subterraneum ssp. brachycalicinum (20.0%) was the highest and weed infestation was half reduced as compared to pure swards. Probably a weaker development of subterranean clover in mixtures with tall fescue was due to the greater competiveness of tall fescue because of its alellopathic potential (Siegel and Bush, 1997; Renne et al., 2004; Marinov-Serafimov et al., 2015).

Regarding the part of other two subspecies-there was the same tendency as about the cocksfoot. Trifolium subterraneum ssp. yaninicum and Trifolium subterraneum ssp. subterraneum parts were 11.0 and 5.3%, respectively and up 18.0% for the clovers in the four-component mixture.

There were no weed infestation in the second cut. Tall fescue is aggressive against the weeds too. Part of clovers was 12.2, 10.4 and 5.8% for Trifolium subterraneum ssp. brachycalicinum, Trifolium subterraneum ssp. yaninicum and Trifolium subterraneum ssp. subterraneum. This part in the four-component mixture was 14.3%.

Pecetti and Piano (1998) consider that in subterranean clover, leaf size contributes to plant competitively. In our study the leaf/stem ratio (fresh weight) of grass components were calculated. This ratio is an important factor affecting quality and forage intake (Smart et al., 1998; Ostrem et al., 2013).

Leaf/stem ratio in the year after sowing was lower due to slowly development of grasses. For cocksfoot leaf/stem ratio varied from 0.44-0.59 and for tall fescue from 0.28-0.36 (Fig. 5). Similar were Shaffer et al. (1994) findings for tall fescue.

The values of this ratio were higher in mixtures as compared to pure grasses.

Fig. 5: Leaf/stem ratio of cocksfoot and tall fescue pure and in mixture with subterranean clover

More leaf biomass formation from grasses in mixtures with legume component was associated to better nitrogen assimilation (Ledgard and Steel, 1992).


The inclusion of subterranean clover as a component of mixtures with cocksfoot and tall fescue improved botanical composition and decreased the weed infestation in the swards. Trifolium subterraneum ssp. brachycalicinum in mixture with cocksfoot had the biggest part in the sward (31.0%) and reduced the weeds half (43.9%) in the year after sowing. Trifolium subterraneum ssp. brachycalicinum in mixtures with tall fescue had small part and decreased the weeds by 28.1%. Subterranean clover as a legume component contributed to increasing the leaf/stem ratio of cocksfoot and tall fescue and it was higher as compared to this ratio in pure grown grasses.

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