Field Crops Research 155 (2014) 67–76

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Field Crops Research

journal homepage: www.elsevier.com/locate/fcr

Crop growth, light utilization and yield of relay intercropped cotton as

affected by plant density and a plant growth regulator
Lili Mao a , Lizhen Zhang b,e,∗ , Xinhua Zhao c , Shaodong Liu c , Wopke van der Werf d ,
Siping Zhang c , Huub Spiertz d , Zhaohu Li a,e
a
China Agricultural University, College of Agronomy and Biotechnology, Beijing 100193, China
b
China Agricultural University, College of Resources and Environmental Sciences, Beijing 100193, China
c
Institute of Cotton Research of Chinese Academy of Agricultural Sciences, Key Laboratory of Cotton Genetic Improvement, Anyang, Henan 455004, China
d
Wageningen University, Centre for Crop Systems Analysis, Crop & Weed Ecology Group, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands
e
Hebei Cottonseed Engineering Technology Research Center, Hejian 062450, Hebei, China

a r t i c l e i n f o a b s t r a c t

Article history: Modern cotton cultivation requires high plant densities and compact plants. Here we study planting
Received 26 August 2013 density and growth regulator effects on plant structure and production of cotton when the cotton is grown
Received in revised form in a relay intercrop with wheat, a cultivation system that is widespread in China. Field experiments were
24 September 2013
carried out in 2010, 2011 and 2012 in Anyang, Henan province, China. Plant densities (PD) were 3.0, 4.5, 6.0
Accepted 25 September 2013
and 7.5 plants m−2 , and growth regulator mepiquat chloride (MC) was applied in four different schedules.
Plant density significantly affected cotton biomass, but MC did not. Aboveground biomass was linearly
Keywords:
associated with plant density. Increasing plant density significantly increased crop light use efficiency,
Biomass
Canopy structure
especially during the reproductive phase. This effect was attributed to a better light distribution in the
Light interception canopy, resulting in higher crop photosynthesis. MC increased the partitioning to leaves, expressed as
Light use efficiency (LUE) leaf/shoot ratio. Plant height and length of fruit branches were significantly reduced by MC, resulting in a
Mepiquat chloride more compact canopy. Maximum leaf area index was slightly lowered at higher MC dose, but MC did not
Relay-intercropping significantly affect light interception. Plant density and MC showed a significant interaction effect on crop
height, but not on leaf growth, biomass or lint yield. At high plant densities, 3–4 consecutive applications
of MC improved plant architecture, resulting in a higher LUE and yield. Lint yields were about 10% higher
with MC applied at a high cumulative dose with high plant densities compared to MC free control.
© 2013 Elsevier B.V. All rights reserved.

1. Introduction Mepiquat chloride (MC) has been reported to modify cotton

Cotton growth is indeterminate. Vegetative and reproductive
growth occur simultaneously (Oosterhuis, 2001). Under optimum
growing condition, plants often become too tall and leafy resulting
in excessively shading (Eaton, 1955) and shedding of reproductive
parts. Much labor is required to cut the top buds of main stem
(cutout) and branches (cutside) to terminate excessive growth. The
growth regulator Mepiquat chloride (N,N-dimethylpiperidinium),
a gibberellin biosynthesis inhibitor (Rademacher, 2000), is widely
used in cotton to control excessive vegetative growth (Ren et al.,
2013).
∗ Corresponding author at: China Agricultural University, College of Resources
and Environmental Sciences, Beijing 100193, China. Tel.: +86 10 62731423;
fax: +86 10 62731423.
E-mail addresses: maolili6666@163.com (L. Mao), Zhanglizhen@cau.edu.cn,
zhang.lizhen@hotmail.com (L. Zhang).
morphological growth, such as reducing leaf area (Reddy et al.,
1996), decreasing plant height (Mondino et al., 2004), shortening
internodes (Fernandez et al., 1991). A more compact plant structure
(Reddy et al., 1990) improves light penetration in the canopy. Appli-
cation of MC decreased the fraction of light intercepted by the upper
leaves (leaves in the top of the canopy), while increasing light pen-
etration to the middle and lower levels of the canopy (Gwathmey
et al., 1995). Multiple applications of MC significantly advanced the
opening of flowers compared with a single application (Biles and
Cothren, 2001). The application of MC decreased shoot length, but
had no significant effect on dry matter production (De Almeida and
Rosolem, 2012). MC concentrated the boll set on lower branches,
increasing the synchrony of boll maturation and supply of assimi-
lates (Gwathmey and Clement, 2010), and increased the fraction of
dry matter allocated to the fruits (Zhao and Oosterhuis, 2000; De
Almeida and Rosolem, 2012).
Gonias et al. (2012) found that light use efficiency (LUE) of cotton
was significantly increased by MC application. Application of MC
increased single leaf photosynthesis (Zhao and Oosterhuis, 2000)

0378-4290/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.fcr.2013.09.021
68 L. Mao et al. / Field Crops Research 155 (2014) 67–76
and canopy CO2 exchange rate (Hodges et al., 1991), in associa-
tion with a higher specific leaf weight (Reddy et al., 1990; Tom
and Oosterhuis, 1993; Zhao and Oosterhuis, 2000). Reddy et al.
(1996) reported that cotton plants developed thicker leaves with
more chlorophyll by MC treatments. A higher net photosynthetic
rate was confirmed in recent studies (Fan et al., 2013; Vermeulen
et al., 2013). A higher specific leaf weight was associated with less
stem growth, reflecting that more carbohydrates accumulated in
the leaves (Reddy et al., 1992). MC changed leaf coloration to darker
green (Gausman et al., 1978) and increased leaf chlorophyll content
(Brand, 1997), which can directly affect photosynthetic potential
and yield (Curran et al., 1990). Thus, there are good indications that
application of MC can increase light use efficiency through an effect
on light distribution in the canopy and increased photosynthetic
rate of leaves.
Cotton canopy structure, light interception and fruit forma-
tion are affected by plant population density (Gwathmey and
Clement, 2010; Kaggwa-Asiimwea et al., 2013). Experiments on
plant spacing have shown that it alters the plant height, archi-
tecture, photosynthetic efficiency of leaves, and boll size (Hussain
et al., 2000; Siebert and Stewart, 2006). Leaf area index (LAI)
and lint yield increase with plant density (PD), but the area of
individual leaves is reduced (Gwathmey and Clement, 2010). Dry
matter partitioning to fruit was higher at 11 plants m−2 than at 6
or 15 plants m−2 (Wang et al., 2011). Niinemets (2010) indicated
change in leaf, shoot and canopy size, determines light harvesting
and the light-driven plasticity, e.g. foliage inclination angle distri-
bution and spatial aggregation. More quantitative studies on cotton
light utilization along plant density gradients are required.
While most of the world’s cotton is grown in single crop sys-
tems, relay intercropping of cotton with winter wheat (Fig. 1) is a
common farming system in the Yellow River region (Zhang et al.,
2007). In this cultivation system, wheat is grown in strips of 3 or
4 rows wide, and harvested in July. Wheat yield per row is higher
than in single wheat cultivation because of greater tillering and
higher yield per plant in the border rows of the wheat strips, where
the plants have more space for growth. Wheat yield per ha in inter-
cropping is approximately 70% of wheat yield in sole crop (Zhang
et al., 2007). Cotton is grown in strips of 1–2 rows in the space that
is left bare between the wheat strips. The sowing is in late April,
and the cotton is in the seedling stage at wheat harvest (Fig. 1A).
Cotton growth is initially delayed due to shading and other com-
petitive influences of wheat (Zhang et al., 2008b), but after wheat
harvest, the expanding cotton canopy will occupy the whole field,
and intercept most of the incoming radiation (Zhang et al., 2008a).
Cotton yield in intercrop is approximately 70% of the cotton yield in
Fig. 1. Photos of wheat-cotton intercrop (A) during the cotton seedling stage (20 June, 5 days after mechanical wheat harvest), and (B) during flowering and boll stage of
cotton on 25 July when the cotton leaf canopy is largely covering the wheat stubble. The picture on the right hand side shows the effect of 4 applications of mepiquat chloride
(M4) as compared to control (M0).
single crop (Zhang et al., 2007). The relative yield total (equal to the
land equivalent ratio) in this system is approximately 1.4, due to the
complementary timing of resource capture of the two crop species,
contributing to high land use efficiency. It is unknown how changes
in plant density of cotton within the strips in the intercrop would
affect canopy structure, light capture and crop light use efficiency.
Cotton growth, canopy structure and light utilization can be
enhanced by optimizing MC dosing and timing in the intercrops.
Such optimization of MC use should be coordinated with the effects
of plant density in the intercrop context. However, limited knowl-
edge of the underlying processes is available. Therefore, objectives
of the study were to quantify the effects of MC and PD on: (a) growth
and dry matter accumulation of cotton in a row-structured inter-
crop; (b) plant morphology dynamics, e.g. plant height, length of
fruit branches, in relation to plant density and MC application, to
determine cotton canopy light capture; and (c) light interception
and use efficiency in response to plant density and MC.
2. Materials and methods
2.1. Field experiments
Field experiments were conducted from 2010 to 2012 in Anyang
city, Henan province, China (36◦ 07 N, 116◦ 22 E), located in the Yel-
low River cotton producing region. Soil type is a sandy loam, with
a pH of 8.0, total N of 1.02 g kg−1 , total P of 0.52 g kg−1 , total K of
17.3 g kg−1 , organic matter content of 13.2 g kg−1 , and a soil bulk
density of 1.36 g cm−3 (averaged over the top 100 cm). Monthly
mean temperatures and totals of rainfall and sunshine hours in the
three years are listed in Table 1. Cotton (Gossypium hirsutum) was
relay intercropped with winter wheat (Triticum aestivum). Wheat
was sown in November of the previous year after cotton was har-
vested. The field experiments were carried out as a randomized
complete block design with 4 plant densities, 5 MC treatments and
3 replicates. Each plot was 33.6 m2 (4.2 m width × 8.0 m length).
The four cotton plant densities were 3.0, 4.5, 6.0, and 7.5 plants m−2 .
The five MC treatments, coded as M0 through M4, included an MC-
free control (M0), a single MC application at the boll stage (M1),
two applications, viz. at flowering and boll stage (M2), three appli-
cations, viz. at squaring, flowering and boll stage (M3), and four
applications, viz. at seedling, squaring, flowering and boll stage
(M4). Treatments doses were tailored to each plant stage accord-
ing to expert knowledge: 6.0, 18.0, 45.0 and 60.0 g ha−1 at seedling,
squaring, flowering and boll stage, respectively. The total doses for
M1-M4 amounted to 60, 105, 123, and 129 g ha−1 (Table 2). MC was
applied by a dorsal manual sprayer.
 L. Mao et al. / Field Crops Research 155 (2014) 67–76 69

Table 1
Monthly weather data during three cotton growing seasons (2010–2012) at the experimental site near Anyang, China.

Average temperature (◦ C) Rainfall (mm) Sunshine hours (h)

2010 2011 2012 2010 2011 2012 2010 2011 2012

April 12.0 14.8 16.8 19.6 7.6 45.6 186 238 175
May 20.8 21.0 22.7 57.0 41.2 4.9 214 213 190
June 26.3 27.0 26.8 20.2 13.5 44.7 211 188 168
July 28.0 27.3 27.9 89.4 84.0 184.1 184 177 133
August 24.8 24.6 25.0 223.8 152.6 145.9 126 111 112
September 20.6 18.2 20.9 111.5 109.9 29.3 114 89 160
October 14.3 14.9 16.0 7.8 37.2 12.5 129 150 154
Totala 21.0 21.1 22.3 529.3 446.0 467 1164 1165 1092
a
Indicates the temperature is daily averaged and others are total amounts.

The intercropping system consisted of an alternation between 2
rows cotton and 3 rows of wheat with 40 cm row width for cotton,
20 cm row width for wheat, and 30 cm between wheat and cotton
adjacent rows, resulting in replacement design with a total strip
width (wheat + cotton) of 1.4 m. Wheat was sown on 3 November
2009, 5 November, 2010 and 5 November, 2011, and cotton on 30
April 2010, 29 April 2011 and 29 April 2012, respectively. Wheat
was harvested on 15 June in each year, and cotton bolls were hand
picked at three times during October in each year. The wheat culti-
var was ‘Zhongyu 5’ and the cotton cultivar Bt (Bacillus thuringiensis)
transgenic cotton ‘Guoxin 3’.
All plots received as fertilizer 225 kg ha−1 N, 150 kg ha−1 P2 O5
and 225 kg ha−1 K2 O according to farmer’s practice. Irrigation was
given by flooding the field, based on crop water requirement. In
2010, a total 222 mm was delivered: 44 mm on 7 March, 39 mm on 2
April, 52 mm on 22 April, 87 mm on 24 June. In 2011 a total amount
of 200 mm of irrigation was given: 34 mm on 23 February, 35 mm
on 31 March, 44 mm on 23 April, 61 mm on 20 June and 26 mm on
26 July. In 2012, 180 mm was applied: 63 mm on 10 March, 67 mm
on 16 April and 50 mm on 11 June.
2.2. Sampling and measurements
Total dry matter (DM), was determined by taking samples per
plot at successive stages: 10 cotton plants at seedling stage and 3
plants from squaring to boll stage. The samples were subdivided
into leaves, stem, squares, flowers and bolls. First, samples were
used for counting the number of organs, measuring plant height,
branch length and all leaf area by using a ruler, and next oven-dried
at 105 ◦ C for 3 h and subsequently at 65 ◦ C to constant weight.
The area of individual leaves was calculated from measurements
as length × width × 0.83 (Zhang et al., 2008a). Leaf area index (LAI)
was calculated by multiplying leaf area per plant with actual plant
density.
The leaf/shoot ratio (LSR) was calculated as the amount of leaf
DM divided by total DM of above-ground vegetative organs. Specific
leaf weight (SLW) was calculated as leaf DM per unit leaf area. LSR
and SLW were measured during the cotton flowering period, when
LAI was at its maximum.
2.3. Calculation of light interception and light use efficiency
Light interception of intercropped cotton was derived from
daily incoming radiation and the calculated fraction of intercepted
radiation in row-structured canopies (Goudriaan, 1977). Incoming
photosynthetically active radiation (PAR), was computed as 50%
of daily global solar radiation, which was derived from sunshine
hours, using Angstrom’s equation with coefficients applicable to
China (Zhou et al., 2005). The daily fraction of intercepted PAR for
intercropping cotton was calculated with a light interception model
developed for wheat-cotton intercrop (Zhang et al., 2008a), using
as inputs measurements of LAI and plant height. The weather data
were collected from an automatic weather station (Campbell Sci.,
Logan, UT) near the experimental field. Light use efficiency (LUE)
was calculated by regressing dry matter increase on intercepted
PAR.
2.4. Data analysis
All statistical analyses were performed by using the Univariate
General Linear Models procedure of SPSS 20. In the analysis, MC, PD,
and year were entered as fixed effects, while block (replicate) was
entered as random factors. The factor block was nested within year.
Least significant differences (LSD) were used to separate treatment
means at the 5% level.
3. Results
3.1. Above-ground biomass and lint yield
Across three years, the above-ground biomass dry weight (DM)
was significantly (P < 0.01) affected by plant density and year, as
shown by significant main effects for these factors (Table 3). There
was also a significant influence of MC, which was not expressed as a
main effect, but only in a significant two-way interaction with year,
and in a significant three-way interaction with year and plant den-
sity, indicating that the effect of MC is modulated by environmental
and management factors affecting the growth and physiology of
the crop. Above ground dry matter showed good positive linear

Table 2
Timing and dose of mepiquat chloride (MC) in intercropped cotton.

Treatment code Number of MC applications MC dose in four development stages Total dose
a
Seedling Squaring Flowering Boll
g ha−1 g ha−1 g ha−1 g ha−1 g ha−1

M0 0 0 0 0 0 0
M1 1 0 0 0 60 60
M2 2 0 0 45 60 105
M3 3 0 18 45 60 123
M4 4 6 18 45 60 129
a
Seedling stage refers to 4–5 main stem nodes; squaring stage refers to 10–11 main stem nodes; flowering stage refers to 15–17 main stem nodes; boll stage refers to one
week after cutout (excision of the growing tip of the main stem in order to terminate its growth).
70 L. Mao et al. / Field Crops Research 155 (2014) 67–76

Table 3
Results of ANOVA on the effects of mepiquat chloride (MC), plant density (PD), year (Y) and their interactions on dry matter (DM), lint yield, plant height, leaf/shoot ratio
(LSR), specific leaf weight (SLW), maximum leaf area index (LAImax ), light interception (LI) and light use efficiency (LUE) in cotton intercropped with wheat from 2010 to
2012 (Anyang, China).

Effect DM Lint yield Height LSR SLW LAImax LI LUE df

Y 60.2** 57.4** 53.9** 43.1** 100.3** 10.4* 81.1** 380.6** 2

MC 1.70ns 5.99** 22.5** 12.0** 0.78ns 4.64** 2.40ns 2.84* 4
PD 126.4** 17.0** 21.7** 0.33ns 9.37** 129.4** 69.5** 75.0** 3
Y × MC 2.99** 1.67 ns 4.04** 1.96ns 2.06* 1.29ns 0.82ns 2.25* 8
Y × PD 8.2** 4.58** 5.09** 2.66* 1.80ns 2.17ns 3.41** 9.90** 6
MC × PD 0.89ns 0.68 ns 2.78** 1.43ns 0.94ns 0.58ns 0.64ns 1.17ns 12
Y × MC × PD 2.57** 0.84 ns 1.81* 1.22ns 2.12** 1.78* 1.22ns 2.10** 24

F values and significance levels (** P < 0.01, * P < 0.05 and ns P0.05) are given.

association (R2 = 0.64–0.85) with plant density when data from dif- 2012 (Fig. 2C). Growing conditions in different years significantly
ferent MC treatments were combined (Fig. 2A–C). MC effects on affected biomass. Both the main effect of year and the two and
DM were significant (P = 0.05) only at the highest plant density three-way interactions of year with PD and MC were significant
(7.5 plants m−2 ), especially for M3 (P = 0.005) and M2 (P = 0.041) in (Table 3).

1200 1600
A 2010 D 2010
Above ground dry matter (g m )
-2

1400
1000

Liny yield (kg ha )

-1 1200
800
1000
600
800
y = 66.926x + 465.85 y = 14.461x + 1010.4
400 2 2
R = 0.7423 600 R = 0.0534

200 400

1600 1600

1400 B 2011 E 2011

Above ground dry matter (g m )
-2

1400

1200
Lint yield (kg ha )
-1

1200
1000
1000
800
800
600
y = 84.457x + 461.51 y = 60.984x + 705.58
400 2
R = 0.6389 600 2
R = 0.4538

200 400

1800 2000

1600 C 2012 F 2012

1800
Above ground dry matter (g m )
-2

1400
1600
Lint yield (kg ha )
-1

1200
1400
1000
1200
800
1000
600 y = 139.45x + 275.25 y = 73.028x + 1168.7
2 2
400 R = 0.8492 800 R = 0.5905

200 600
2 3 4 5 6 7 8 2 3 4 5 6 7 8
-2 Plant density (plants m-2)
Plant density (plants m )
M0 M1 M2 M3 M4 M0 M1 M2 M3 M4

Fig. 2. Relationship between above ground dry matter and plant density (A–C), and between lint yield and plant density (D–F) in three years (2010–2012) in four different
MC treatments (M1–M4: 1–4 applications of MC) and control (M0) (cf. Table 2).
 L. Mao et al. / Field Crops Research 155 (2014) 67–76 71

120 70
a
A B
100 a 60 a

Length of fruit branch (cm)

a a ab
ab b b b b
b 50
80

Plant height (cm)

b a
40
60 b
b a
30
40
20 b
20 10
0 0
3 4.5 6 7.5 Fruit branches in Fruit branches in Fruit branches in
Plant density (plants m-2) low position middle position upper position
M0 M3 M4 (1-4 fruit (5-9 fruit (>10 fruit
branches) branches) branches)

Fig. 3. Plant height (A) and fruiting branch length for 3.0 plants m−2 (B) of intercropped cotton as affected by mepiquat chloride (MC) in 2011 and 2012 (Anyang, China). M3
and M4 indicate MC treatments with 3 or 4 applications, respectively (cf. Table 2). M0 indicates MC free control.

Lint yield was significantly affected by MC, plant density and 0.6
year, but the interaction of plant density and MC was not significant,

Leaf/shoot ratio (g g ) ,
indicating consistency of MC effect over plant densities (Table 3). 0.5 a a a

-1
ab a a a
Lint yields were 1600 kg ha−1 at the highest plant density in the a a a
bc c bc a ab
ab

warmest year (2012) and increased linearly with plant density in 0.4 ab a b
b
2011 and 2012 (Fig. 2D and E), but there was no significant increase
0.3
of lint yield with plant density in 2010 (Fig. 2D; P = 0.364). In the
wet year of 2010, lint yield increased significantly (P = 0.04) at high 0.2
plant densities (6.0 and 7.5 plants m−2 ) with MC applied at four
times (M4), on average by 11%, as compared to the control (Fig. 2D). 0.1

3.2. Canopy structure 0

3 4.5 6 7.5
-2
Final plant height was significantly affected by interactions Plant density (plants m )
between PD, MC and year (Table 3). At a high plant density of M0 M1 M2 M3 M4
7.5 plants m−2 , plant height was significantly reduced in the heavi-
est MC treatments M4 and M3, as compared to the control (Fig. 3A). Fig. 4. Leaf/shoot ratio of intercropped cotton in relation to plant density and appli-
There was a tendency toward greater plant height at higher plant cation of mepiquat chloride (MC) in 2011 (Anyang, China). The codes M0-M4 indicate
0–4 applications of MC (cf. Table 2).
density in control treatments, but this tendency was not significant
(Fig. 3A); however, height was not significantly affected by density
if MC was applied (Fig. 3A).
Fruit branch length was significantly shortened (30–50%) by MC
treatment M4 (Fig. 3B), especially at lower positions (1–4 fruit
branches). Thus, a more compact crop canopy was achieved by
applying MC due to significant decrease of plant height and length
of fruit branches. 90
85
3.3. Leaf morphology and LAI
Specific leaf weight (g m ) ,

80
-2

Leaf/shoot ratio (LSR) was significantly (P < 0.01) affected by MC 75

and year, but it was not affected by plant density (Table 3). There
70
was a tendency to an increase in LSR with MC dose, especially in
highest plant density (Fig. 4). 65
Specific leaf weight was significantly affected by plant den- 60
sity but not by MC. SLW decreased with a rise in PD from 3
to 6 plants m−2 , but did not change at PD above 6 plants m−2 55
(Fig. 5). However, SLWs in high plant densities (6 plants m−2 and 50 M0
7.5 plants m−2 ) were significantly higher with MC (Fig. 5). The effect
of MC significantly interacted with year (P < 0.05).
45 M4
Leaf area index was affected both by PD and MC. LAI values 40
at 100–130 days after sowing (DAS) were significantly (P = 0.002)
2 3 4 5 6 7 8
reduced by MC (Fig. 6). LAImax varied with plant density, from -2
about 2.5 to 5.0. The effects of PD and MC on LAImax were additive
Plant density (plants m )
(Table 3), and the reduction of LAI by MC was readily compensated Fig. 5. Specific leaf weight of intercropped cotton in relation to plant density and
at higher plant density (Fig. 6D). Overall, the effect of plant density application of mepiquat chloride (MC) in 2011 (Anyang, China). M0: MC-free control.
on LAI was substantially greater than that of MC. M4: four applications of MC (cf. Table 2).
72 L. Mao et al. / Field Crops Research 155 (2014) 67–76

6
A 3.0 plants m-2 B 4.5 plants m-2
5
Leaf area index
4

0
6
C 6.0 plants m-2 D 7. 5 plants m-2
5
Leaf area index

2
M0
1
M4

0
0 20 40 60 80 100 120 140 160 0 20 40 60 80 100 120 140 160
Days after sowing (d) Days after sowing (d)

Fig. 6. Leaf area index (LAI) of intercropped cotton as affected by mepiquat chloride (MC) at different plant densities in 2011 (Anyang, China). M0 is the MC-free control, and
M4 is the treatment with four MC applications (cf. Table 2).

3.4. Light interception (LI) and Light use efficiency (LUE)
Light interception was significantly (P < 0.01) affected by plant
density (Table 3), showing an increase with plant densities (Fig. 7
D-F). Although MC slightly reduced LAI, light interception (LI) was
not significantly different between MC treatments (Table 3). Con-
cerning LI, MC treatments showed no significant interactions with
plant density and year, while the interaction between PD and year
was significant (Table 3).
Light use efficiency (LUE), the slope of the linear relationship
between above-ground DM and total light (PAR) interception, was
significantly affected by MC, PD and year (Table 3). The overall
LUE for all treatments in 2010, a wet year, amounted to 1.54 g
DM MJ PAR−1 , about 32% lower than in the drier years of 2011 and
2012: 2.24 g DM MJ PAR−1 (Table 4). The total of sunshine hours was
similar in three experimental years, but rainfall during the cotton
growing season in 2010 was 16.0% higher than in 2011 and 2012.
LUE, averaged over MC treatments, was 2.32 DM MJ PAR−1 at the
highest plant density vs. 1.78 g DM MJ−1 PAR at the lowest plant
density, a difference of 30% (Table 4). Increasing LUE at high plant
density was due to the decrease of DM (Fig. 7A–C) at late cotton
growing season (after 139 DAS), especially in drier years of 2011
and 2012, without the decrease of light interception rates in this
season (Fig. 7D–F).

Table 4
Light use efficiency (g DM MJ PAR−1 ) of intercropped cotton in relation to plant density and application of mepiquat chloride (MC). M0-M4 indicate MC treatments (cf. Table 2).

MC code Plant density (plants m-2 ) Mean

3.0 4.5 6.0 7.5

2010 2011 2012 2010 2011 2012 2010 2011 2012 2010 2011 2012

M0 1.26 a 2.05 a 1.84 a 1.58 a 1.93 b 1.90 a 1.63 a 2.34 a 2.56 a 1.80 ab 2.60 a 2.42 b 1.99 b
M1 1.22 a 2.20 a 1.82 a 1.28 ab 2.31 ab 2.01 a 1.73 a 2.55 a 2.29 a 1.50 c 2.62 a 2.74 ab 2.02 b
M2 1.35 a 2.41 a 1.60 b 1.26 b 2.06 ab 1.96 a 1.46 b 2.23 a 2.23 a 1.95 a 2.45 ab 2.75 ab 1.98 b
M3 1.39 a 2.38 a 1.89 a 1.57 ab 2.34 a 2.13 a 1.75 a 2.38 a 2.62 a 1.71 b 2.14 b 3.09 a 2.12 a
M4 1.39 a 2.13 a 1.81 ab 1.55 ab 2.05 ab 2.07 a 1.72 a 2.47 a 2.38 a 1.73 b 2.59 a 2.68 ab 2.05 ab
SE 0.08 0.12 0.07 0.10 0.11 0.10 0.05 0.17 0.13 0.05 0.15 0.15 0.03
Mean ± SE 1.78 ± 0.061 1.87 ± 0.055 2.16 ± 0.063 2.32 ± 0.075 2.03 ± 0.036

Small letter indicates no significance at 0.05 level.

 L. Mao et al. / Field Crops Research 155 (2014) 67–76 73

1400
A 2010 B 2011 C 2012
1200
Dry matter (g m-2) 1000
800
600
400
200
0

500
D 2010 E 2011 F 2012
Light interception (MJ m-2)

400

300

200

100

0
0 50 100 150 200 0 50 100 150 200 0 50 100 150 200
Days after sowing (d) Days after sowing (d) Days after sowing (d)

3.0pla
M0, 3.0 pla
ntnts
m-2m-2 3.0pla
M4, 3.0 pla
ntnts
m-2m-2 7.5pla
M0, 7.5 pla
ntnts
m-2m-2 7.5pla
M4, 7.5 pla
ntnts
m-2m-2

Fig. 7. Total dry matter accumulation (A–C) and cumulative light interception (D–F) of intercropped cotton as affected by plant density and mepiquat chloride (MC) from
2010 to 2012. M4 is the MC treatment with 4 applications. M0 is MC free control (cf. Table 2).

Averaged over years, applying MC from cotton squaring stage Applying MC (M4 and M3) at an early growth stage (DAS = 80),
onwards increased LUE by 6.5%; from 1.99 g MJ PAR−1 in the showed a stronger effect on LUE at low plant densities (Fig. 8A)
control to 2.12 g MJ PAR−1 in M3 (Table 4). Applying MC from the compared to treatments without MC at this stage (M0). However,
seedling stage onwards (M4) resulted in a slight and insignificant LUE was not affected by MC at a high plant density (Fig. 8B), which
increase of LUE, while applying MC from flowering and boll stages indicates that the MC effect may have been suppressed at high
onwards showed no difference compared to the control (Table 4). plant densities. The significant increase of LUE late in the season
Thus timing of MC affects the effect of MC on light use efficiency. (after 139 DAS) at higher plant densities for both with and without

2
a
A 3.0 plants m-2 B 7.5 plants m-2 a
Light use efficiency (g MJ PAR-1)

1.8 a
a
1.6 a
a a
a a a
1.4 ab b ab
b
1.2
b
1 a a
a
0.8
0.6
0.4
0.2
0
2010 2011 2012 2010 2011 2012
Year Year
M0 M3 M4

Fig. 8. Light use efficiency during the early cotton growing period (80 days after sowing) in relation to mepiquat chloride (MC). Treatment codes M0, M3 and M4 indicate
the number of MC sprays that were applied (cf. Table 2).
74 L. Mao et al. / Field Crops Research 155 (2014) 67–76
Fig. 9. Light use efficiency at late cotton growing season (from 139 to 149 days after sowing) in relation to mepiquat chloride (MC) and plant density in 2010–2012. M0:
MC-free control. M4: four applications of MC (cf. Table 2).
MC treatments (Fig. 9) were due to a higher growth rate per unit
ground area.
4. Discussion and conclusions
4.1. Biomass and lint yield
The finding that cotton biomass in a relay strip intercropping
system increased linearly with plant density, with no significant
overall main effect of MC, is consistent with results reported by De
Almeida and Rosolem (2012). However, in our study, effects of plant
density and MC on biomass strongly interacted with weather con-
ditions in different years. Thus, the effects of MC may be different
between years according to the weather conditions. Lint yield of
cotton linearly increased with plant density and was only slightly
increased by MC with 4 applications at high plant densities in a wet
year. High plant densities reduced boll weight, but increased boll
numbers per unit area, and therefore increased lint yield in short-
season cotton (Dong et al., 2010). High densities increase yields by
raising boll numbers (Bednarz et al., 2006), inhibiting the occur-
rence of outer bolls, and improving boll distribution (Gwathmey
and Clement, 2010). However, there was no significant increase
in lint yield at higher plant density (Bednarz et al., 2000; Dong
et al., 2006). MC is considered to adjust vegetative and reproductive
growth, resulting in improved cotton yields (Oosterhuis and Egilla,
1996). It was also reported that MC increases the fraction of bolls
set on the first five fruiting branches, and decreases the fraction of
bolls above top fruiting branches (Gwathmey and Clement, 2010;
Zhao and Oosterhuis, 2000), but does not alter the horizontal boll
distribution. Ren et al. (2013) reported that MC increased single boll
weight. Our findings showed that the effects of plant growth reg-
ulator on lint yield depend on the weather. The best results of MC
were found at the highest density in a wet year. Successive appli-
cations of a plant growth regulator in cotton may be an investment
to avoid yield losses in years with rainy weather. Saving on the
costs of the use of plant growth regulator could be made possible
by optimizing plant densities and/or by selection of cultivars better
adapted to high densities.
4.2. Plant morphology and canopy structure
With respect to plant morphology of cotton in intercropping
systems, plant height and fruit branch length were reduced by
MC treatments, especially at high plant densities. The increasing
trend of plant height associated with plant density was significantly
suppressed by applying MC. A similar plant height reduction by
plant growth regulator applications was reported by Pettigrew and
Johnson (2005). York (1983) found that higher plant densities led
to cotton crops with excessive vegetative growth that was more
responsive to MC. The reduction of plant height is effective to opti-
mize plant height at high plant densities for mechanical harvesting
(Oz et al., 2011).
Intercropping itself does also change canopy architecture. The
number of leaves and fruit branches shows a significant decrease
compared to sole cotton, but fruit node number increases under MC
free conditions (Zhang et al., 2008b). MC and PD effects on mor-
phology in intercrops may differ from those in sole cotton. Thus,
findings in intercropping cannot be extrapolated to crops grown in
monoculture without experimental validation.
4.3. Light interception and use efficiencies
Light interception of intercropped cotton was significantly
increased by higher plant density in association with a strong
increase of LAI. This was partially due to more leaves, and partially
to thinner and larger leaves. Light interception was not decreased
by MC treatments, because LAI was only slightly reduced. The light
extinction coefficient may be reduced by more erect leaves (Gu
et al., 2013). It has been reported that MC results in shorter and
more compact plant architecture, because of reduced stem elonga-
tion, leaf expansion, and leaf area (Reddy et al., 1990). We found
that light use efficiency of intercropped cotton was increased sig-
nificantly by a rise in plant density: 30% higher at a plant density of
7.5 plants m−2 than at 3.0 plants m−2 averaged for all years and MC
treatments. A significant increase of LUE at higher plant density was
also found in pigeon pea in southern Ethiopia (Worku and Demisie,
2012) and in wheat grown in a semi-arid area (Wajid et al., 2004).
However, our findings are not consistent with a study of Siebert
and Stewart (2006), who reported that a rise in plant height due
to an increase of plant density leads to more mutual shading at
higher plant densities and thus reduces light utilization efficiency.
The latter is contrary to expectation, however, because at lower
light levels the photosynthetic efficiency will be higher.
Zhang et al. (2008a) found that intercropping had only a slight
effect on light interception of intercropped cotton (93% as much
PAR as in a sole crop). The slight decrease of light interception in
the intercropping system might not only be due to the strip struc-
ture, but may also have been caused by a pronounced delay in early
 L. Mao et al. / Field Crops Research 155 (2014) 67–76
development (Zhang et al., 2008b). The LUE of monoculture was
1.39 g DM MJ PAR−1 and 1.33 g DM MJ PAR−1 for similar intercrop-
ping (Zhang et al., 2008a). This value is close to LUE in our study
(1.78 g DM MJ PAR−1 ) for similar plant density.
We conclude the mechanism for increasing LUE could be largely
determined by the indirect effects of a change in morphological
traits due to effects of plant density and MC on the crop physiol-
ogy. With respect to the effect of increased plant density on LUE, this
may be due to more growth at late season by an increased number
of green leaves in the canopy. Furthermore, a delay of leaf senes-
cence at higher density is suggested by Dong et al. (2012). We also
assumed that the rise in LAI at higher plant density did not increase
the canopy cover ratio due to the already high light extinction coef-
ficient (0.95–0.98; Zhang et al., 2008a). Thus, total light interception
does not correspondingly increase with a rise in biomass, especially,
after canopy closure. Another explanation may be that modifica-
tion of light extinction coefficient resulting from plant branching
and leaf angles by intercropping and MC (Gu et al., 2013), probably
also is modified by plant density. Those assumptions need to be
studied in more detail.
Light use efficiency was only slightly increased by MC in this
study, while it was significantly increased by three applications of
MC (Gonias et al., 2012). It was mainly due to the modification of
plant structure e.g. plant height and leaf angles (Gu et al., 2013),
increment of leaf thickness in Reddy et al. (1996) and physiological
traits, e.g. chlorophyll content (Reddy et al., 1996; Rethwisch et al.,
2010).
4.4. Optimizing plant density and MC applications
Our results showed that the interactions between MC and plant
density on biomass and lint yield, LAI, light interception and use
efficiency were affected by seasonal effects; on average those were
not significant, but the interaction between plant density, MC and
year was significant. Similar results were found on lint yield and
yield components (boll number and boll weight) in sole cotton
experiments, which might be due to sub-optimal plant densities
(Ren et al., 2013). We conclude that the effects of MC and plant
density may be compensatory: the negative effects on cotton struc-
tures due to PD, e.g. excessive increase of LAI, plant height and
length fruit branches, could partially be compensated by an effect
of MC on canopy size (more compact) and a reduction of LAI. Higher
plant densities will increase intra-plant competition for resources,
e.g. light, water and nutrients; however, MC does make the canopy
more compact and could therefore decrease the competition for
light. It implies that optimizing plant density combined with MC
treatments could achieve an optimal canopy that favors light inter-
ception and utilization.
We hypothesize that here was still a potential for significant
interactions of MC and plant density to improve yield and light
use efficiency, if we had imposed higher plant densities in this
study (see also: Ren et al., 2013). Optimizing plant density and
MC for improving lint yield and resource use efficiency in cotton is
complex, because of the interactions and trade-offs between mor-
phological and physiological processes during the growth cycle. The
mechanisms of functional responses to changes in structural mor-
phological traits, e.g. canopy size, branch orientation, are difficult
to quantify in field experiments. Questions to address are: a. What
are the mechanisms that are responsible for higher light use effi-
ciency at higher plant density? b. What is the role of crop geometry
in relation to vertical light distribution? These questions may be
addressed by functional-structural plant modeling (FSPM) as has
been developed by Evers et al. (2011). A better quantitative insight
in mechanistic processes will be helpful to explore effectiveness of
improved management options.
75
Acknowledgements
This research was supported by Transgenic major project
(2012ZX08013010), ‘948’ Program (2011-G19) and the pro-
gram of the Modern Agricultural Industry Technology System
(CARS-18-18).
References
Bednarz, C.W., Bridges, D.C., Brown, S.M., 2000. Analysis of cotton yield stability
across population densities. Agron. J. 92, 128–135.
Bednarz, C.W., Nichols, R.L., Brown, S.M., 2006. Plant density modifications of cotton
within-boll yield components. Crop Sci. 46 (5), 2076–2080.
Biles, S.P., Cothren, J.T., 2001. Flowering and yield response of cotton to application
of mepiquat chloride and PGR-IV. Crop Sci. 41, 1834–1837.
Brand, M.H., 1997. Shade influences plant growth, leaf color and chlorophyll content
of Kalmia latifolia L. cultivars. Hortscience 32, 206–208.
Curran, P.J., Dungan, J.L., Gholz, H.L., 1990. Exploring the relationship between
reflectance red edge and chlorophyll content in slash pine. Tree Physiol. 7, 33–48.
De Almeida, A.Q., Rosolem, C.A., 2012. Cotton root and shoot growth as affected by
application of mepiquat chloride to cotton seeds. Acta Sci. Agron. 34 (1), 61–65.
Dong, H., Li, W., Tang, W., Li.F Z., Zhang, D., Niu, Y., 2006. Yield, quality and leaf
senescence of cotton grown at varying planting dates and plant densities in the
Yellow River Valley of China. Field Crops Res. 98, 106–115.
Dong, H., Li, W., Xin, C., Tang, W., Zhang, D., 2010. Late planting of short-season
cotton in saline fields of the Yellow River Delta. Crop Sci. 50, 292–300.
Dong, H., Li, W., Eneji, A.E., Zhang, D., 2012. Nitrogen rate and plant density effects
on yield and late-season leaf senescence of cotton raised on a saline field. Field
Crops Res. 126, 137–144.
Eaton, F.M., 1955. Physiology of the cotton plant. Annu. Rev. Plant Physiol. 6, 299.
Evers, J.B., van der Krol, A.R., Vos, J., Struik, P.C., 2011. Understanding shoot branching
by modelling form and function. Trends Plant Sci. 16, 464–467.
Fan, X.X., Xu, Z.G., Liu, X.Y., Tang, C.M., Wang, L.W., Han, X.L., 2013. Effects of light
intensity on the growth and leaf development of young tomato plants grown
under a combination of red and blue light. Sci. Horticult. 153, 50–55.
Fernandez, C.J., Cothren, J.T., McInnes, K.J., 1991. Partitioning of biomass in well-
watered and water-stressed cotton plants treated with mepiquat chloride. Crop
Sci. 31, 1224–1228.
Gausman, H.W., Rittig, F.R., Namken, L.N., Rodriguez, R.R., Escobar, D.E., Garza, M.V.,
1978. Effect of 1,1-dimethyl-piperidinium chloride on cotton (Gossypium hir-
sutum L.) leaf chlorophyll, size and structure. In: Proceedings of the 5th Plant
Growth Regulators Working Group Meeting, Blacksburg, Virginia.
Gonias, E.D., Oosterhuis, D.M., Bibi, A.C., 2012. Cotton radiation use efficiency
response to plant growth regulators. J. Agric. Sci. 150, 595–602.
Goudriaan, J., 1977. Crop Micrometerorology: A Simulation Study. Simulation Mono-
graphs. PUDOC, Wageningen, pp. 249.
Gu, S., Evers, J.B., Zhang, L., Mao, L., Vos, J., Li, Z., 8–15 June 2013. Using
functional–structural plant modeling to explore the response of cotton to mepi-
quat chloride application and plant population density. In: 7th International
Conference on Functional-Structural Plant Models (FSPM2013), Saariselkä,
Finland.
Gwathmey, C.O., Clement, J.D., 2010. Alteration of cotton source–sink relations with
plant population density and mepiquat chloride. Field Crops Res. 116, 101–107.
Gwathmey, C.O., Wassel, O.M., Michaud, C.E., 1995. Pix effects on canopy light
interception by contrasting cotton varieties. In: Richter, D.A., Armour, J. (Eds.),
Proceedings of the Beltwide Cotton Conference, 4–7 January 1995, Memphis,
Tennessee. National Cotton Council of America, Memphis, TN, p. 1153.
Hodges, H.F., Reddy, V.R., Reddy, K.R., 1991. Mepiquat chloride and tempera-
ture effects on photosynthesis and respiration of fruiting cotton. Crop Sci. 31,
1302–1308.
Hussain, S.Z., Faird, S., Anwar, M., Gill, M.I., Baugh, M.D., 2000. Effect of plant density
and nitrogen on the yield of seed cotton—variety CIM-443. Sarhad J. Agric. 16
(2), 143–147.
Mondino, M.H., Peterlin, O.A., Garay, F., 2004. Response of late-planted cotton to
application of a growth regulator (chlorocholine chloride, Cycocel). Expt. Agric.
40, 381–387.
Niinemets, U., 2010. A review of light interception in plant stands from leaf to canopy
in different plant functional types and in species with varying shade tolerance.
Ecol. Res. 25, 693–714.
Oosterhuis, D.M., Egilla, J.N., 1996. Field evaluation of plant growth regulators for
effect on the growth and yield of cotton. In: Dugger, P., Richter, D. (Eds.), Proc.
Beltwide Cotton Conf. National Cotton Council, Memphis, TN, pp. 1213–1215.
Oosterhuis, D., 2001. Physiology and nutrition of high yielding cotton in the USA.
Inf. Agron. 95, 18–24.
Oz, E., Tekin, A.B., Evcim, H.U., Degirmencioglu, A., 2011. Effect of variety and row
spacing on the performance of a cotton picker. J. Food Agric. Environ. 9 (1),
236–242.
Pettigrew, W.T., Johnson, J.T., 2005. Effects of different seeding rates and plant
growth regulators on early-planted cotton. J. Cotton Sci. 9, 189–198.
Rademacher, W., 2000. Growth Retardants: effects on gibberellin biosynthesis and
other metabolic pathways. Annu. Rev. Plant Physiol. 51, 501–531.
Reddy, A.R., Reddy, K.R., Hodges, H.F., 1996. Mepiquat chloride (PIX) induced changes
in photosynthesis and growth of cotton. Plant Growth Regul. 20, 179–183.
76 L. Mao et al. / Field Crops Research 155 (2014) 67–76
Reddy, V.R., Baker, D.N., Hodges, H.F., 1990. Temperature and mepiquat chloride
effects on cotton canopy architecture. Agron. J. 82, 190–195.
Reddy, V.R., Trent, A., Acock, B., 1992. Mepiquat chloride and irrigation versus cotton
growth and development. Agron. J. 84, 930–933.
Ren, X., Zhang, L., Du, M., Evers, J.B., van der Werf, W., Tian, X., Li, Z., 2013. Managing
mepiquat chloride and plant density for optimal yield and quality of cotton. Field
Crops Res. 149, 1–10.
Rethwisch, M.D., Dyke, J.V., Williams, M., Grimm, A., Luna, M., 2010. Comparison of
stanceTM and mepiquat chloride plant growth regulators on DPL 164B2RF cotton
in the palo verde valley, 2006. Arizona Cotton Report, March., pp. 159.
Kaggwa-Asiimwea, R., Andrade-Sanchez, P., Wang, G., 2013. Plant architecture influ-
ences growth and yield response of upland cotton to population density. Field
Crops Res. 145, 52–59.
Siebert, J.D., Stewart, A.M., 2006. Influence of plant density on cotton response to
mepiquat chloride application. Agron. J. 98 (6), 1634–1639.
Tom, C.J., Oosterhuis, D.M., 1993. Physiological impact of plant growth regulators.
In: Proceedings of the Beltwide Cotton Production Conference, pp. 128–131.
Vermeulen, P.J., Anten, N.P.R., Stuefer, J.F., During, H.J., 2013. Whole-canopy carbon
gain as a result of selection on individual performance of ten genotypes of a
clonal plant. Oecologia 172, 327–337.
Wajid, A., Hussain, A., Ahmad, A., Rafiq, M., Goheer, A.R., Ibrahim, M., 2004. Effect of
sowing date and plant density on growth, light interception and yield of wheat
under semiarid conditions. Int. J. Agric. Biol. 6 (6), 1119–1123.
Wang, G., Asiimwe, R.K., Andrade, P., August, 2011. Growth and yield response to
plant population of two cotton varieties with different growth habits. Arizona
Cotton Report., pp. 161.
Worku, W., Demisie, W., 2012. Growth, light interception and radiation use effi-
ciency response of pigeon pea (Cajanus cajan) to planting density in southern
Ethiopia. J. Agron. 11 (4), 85–93.
York, A.C., 1983. Response of cotton to mepiquat chloride with varying N rates and
plant populations. Agron. J. 75, 667–672.
Zhao, D., Oosterhuis, D.M., 2000. Pix plus and mepiquat chloride effects on phys-
iology, growth, and yield of field -grown cotton. J. Plant Growth Regul. 19,
415–422.
Zhang, L., van der Werf, W., Zhang, S., Li, B., Spiertz, J.H.J., 2007. Growth, yield and
quality of wheat and cotton in relay strip intercropping systems. Field Crops Res.
103, 178–188.
Zhang, L., van der Werf, W., Bastiaans, L., Zhang, S., Li, B., Spiertz, J.H.J., 2008a. Light
interception and utilization in relay strip intercrops of wheat and cotton. Field
Crops Res. 107, 29–42.
Zhang, L., van der Werf, W., Zhang, S., Li, B., Spiertz, J.H.J., 2008b. Temperature-
mediated developmental delay may limit yield of cotton in relay intercrops with
wheat. Field Crops Res. 106, 258–268.
Zhou, J., Wu, Y.Z., Yan, G., 2005. General formula for estimation of monthly
average daily global solar radiation in China. Energy Conv. Manage. 46,
257–268.