Response of highbush blueberry (Vaccinium

corymbosum) ‘Spartan’ to gibberellic acid and CPPU
applied at various rates and timings
H. Fujisawa, Y. Kawai and K. Ishikawa
Faculty of Agriculture, Tokyo University of Agriculture, Atsugi, Japan.

Abstract
Experiments were conducted in 2010, 2013, 2014, and 2016 at an experimental
field of Tokyo University of Agriculture, Japan, with highbush blueberry (Vaccinium
corymbosum) ‘Spartan’ to determine the effects of varying the concentration and
timing of treatment with gibberellic acid (GA) or CPPU, and to clarify the effects of the
combined application of both growth regulators on fruit size and quality. During the
initial three experimental seasons, GA was applied at 50, 100, and 200 mg L-1 and
CPPU at 5, 10, and 20 mg L-1 on bearing shoots at 0, 5, 10, and 20 days after blooming
(DAB). Both GA and CPPU applications led to increases in fruit size, fruit weight, and
the number of seeds per fruit compared to untreated controls. Fruit produced on
shoots treated with 100 mg L-1 GA was larger than on shoots treated with 50 or 200
mg L-1 GA. GA application was most effective when applied at 5 DAB and was less
effective when applied at 0, 10, or 20 DAB. Higher rates of CPPU application resulted
in larger fruit. However, malformed (wavy) leaves were observed following CPPU
application at 10 and 20 mg L-1. CPPU application was most effective when applied at
10 DAB and was less effective when applied at 0, 5, or 20 DAB. On the basis of these
findings, the combined application of 100 mg L-1 GA at 5 DAB and 10 or 20 mg L-1 CPPU
at 10 DAB was tested in 2014 and 2016. The treatment resulted in a 1.4- to 1.5-fold
increase in mean fruit weight compared to the control, while the soluble solid content
and acidity of the fruit were not significantly affected by the treatment. The combined
application was considerably more effective than application of GA or CPPU alone in
increasing fruit size.

Keywords: acidity, GA, forchlorfenuron, fruit size, fruit weight, number of seeds, soluble
solid content

INTRODUCTION
The fruit of highbush blueberry (Vaccinium corymbosum) produced in Japan for sale in
fresh fruit markets is usually evaluated by the size of the fruit. A larger fruit size is more
profitable to blueberry growers.
Much research has focused on use of the growth regulators gibberellic acid (GA) and
CPPU to enhance fruit production in blueberries. The application of GA was shown to
significantly increase fruit set in highbush blueberry cultivar ‘Coville’ when natural fruit set
was poor; however, the application causes production of smaller fruits (Mainland and Eck,
1969). Similar results were reported following GA application to rabbiteye blueberry
cultivars (Vaccinium ashei) (NeSmith et al., 1995; Cano-Medrano and Darnell, 1998; NeSmith,
2005). By contrast, our previous study demonstrated that GA application to the highbush
blueberry cultivar ‘Spartan’ increased fruit size in parallel with increasing fruit set (Fujisawa
et al., 2011).
The application of the cytokinin-like plant growth regulator CPPU, which is also called
forchlorfenuron, increases fruit size and fruit set in both rabbiteye blueberry (Merino et al.,
2002; NeSmith, 2002, 2008; NeSmith and Adair, 2004) and highbush blueberry (Serri and
Hepp, 2006; Williamson and NeSmith, 2007; Fujisawa et al., 2011; Retamales et al., 2014).
Nevertheless, CPPU does not increase fruit size in the rabbiteye blueberry cultivar ‘Tifblue’
when applied in conjunction with GA (NeSmith, 2002). However, in light of the results from

  Acta Hortic. 1206. ISHS 2018. DOI 10.17660/ActaHortic.2018.1206.25 183

Proc. XIII Intern. Symp. on Plant Bioregulators in Fruit Production
  Ed.: S. Kondo
our previous study (Fujisawa et al., 2011), the highbush blueberry cultivar ‘Spartan’ is
expected to respond positively in terms of fruit size to a combined application of GA and
CPPU.
The objectives of the present study were firstly to determine the appropriate rates and
timings of GA and CPPU application to highbush blueberry ‘Spartan’ as adjudged by the
subsequent fruit size, and secondly to clarify the effects of a combined application of the two
growth regulators on fruit size and quality.

MATERIALS AND METHODS

The experiments were conducted in 2010, 2013, 2014, and 2016 at the experimental
field of the Faculty of Agriculture, Tokyo University of Agriculture, located in Kanagawa, 47
km southwest of Tokyo, Japan. Several northern highbush blueberry cultivars were planted
in a mixed population in the field and allowed to open pollinate.
Mature plants of the cultivar ‘Spartan’ were used in the experiments. Bearing shoots
were selected and sprayed with a solution containing GA (Gibberellin Kyowa,
Kyowahakko-bio Co. Ltd., Tokyo, Japan) or CPPU (Fulmet, Kyowahakko-bio Co. Ltd., Tokyo,
Japan) using a hand sprayer. Control shoots were selected and were untreated. Each
treatment consisted of 4-6 bearing shoots depending on the year.
All the fruit on these shoots was collected and fruit diameter, fruit size distribution
based on fruit diameter, fruit weight, total number of brown and white seeds, soluble solid
content, and titratable acid were measured.

GA application
To investigate the effect of application rate, GA was applied to bearing shoots at 100 or
200 mg L-1 on the day of 50% blooming in 2010 and at 50, 100, or 200 mg L-1 at 5 days after
80% blooming in 2014.
To investigate the effect of application timing, 200 mg L-1 GA was applied on the day of
50% blooming, at 5, 10, or 20 days after blooming (DAB) in 2010. In 2013, 100 mg L-1 GA
was applied on the day of 80% blooming, at 5, or 10 DAB.

CPPU application
To investigate the effect of application rate, 10 or 20 mg L-1 CPPU were applied at 10
days after 50% blooming in 2010 and at 5, 10, or 20 mg L-1 at 10 days after 80% blooming in
2014.
To investigate the effect of application timing, 20 mg L-1 CPPU was applied on the day
of 50% blooming, at 5, 10, or 20 DAB in 2010. In 2013, 10 mg L-1 CPPU was applied on the
day of 80% blooming, at 5, or 10 DAB.

Combined application of GA and CPPU

The combined application of GA and CPPU was tested in 2014 and 2016 in addition to
single applications of GA or CPPU. GA was applied at 100 mg L-1 at 5 days after 80%
blooming. CPPU was applied at 10 or 20 mg L-1 (in 2014 or 2016, respectively) at 10 DAB. In
the combined application treatments, GA application at 5 DAB was followed by CPPU
application at 10 DAB.

RESULTS AND DISCUSSION

GA application
Blueberry fruit harvested from shoots treated with GA were larger, heavier, and had a
higher number of seeds per fruit, regardless of GA rate, than those harvested from untreated
control shoots in both 2010 and 2014 (Table 1). Soluble solid contents were not significantly
different in either year; titratable acid was not significantly different in 2014.
According to previous studies, GA application promotes fruit set due to parthenocarpy
if fruit set is poor; however, GA treatment is accompanied by a reduction in fruit size
(NeSmith et al., 1995; Cano-Medrano and Darnell, 1998; NeSmith, 2005). By contrast, in our

184
experiments, no seedless fruit was observed and the number of seeds per fruit was relatively
high even in the controls. This result implies that GA application increased fruit size and
weight when cross-pollination had occurred. This interpretation corresponds with the
findings in our previous study (Fujisawa et al., 2011).

Table 1. Response of highbush blueberry cultivar ‘Spartan’ to GA applied at various

concentrations.
Fruit size Soluble
GA application Fruit Fruit Titratable
distribution (%) Number of solid
diameter weight acid
Rate Timing <15 15-18 >18 seeds fruit-1 content
(mm) (g) (%)
(mg L-1) (DAB1) (mm) (mm) (mm) (°Brix)
Year 2010
Control 14.1 a2 - - - 1.33 a 20.8 a 13.1 a -
100 0 16.3 b - - - 1.94 b 44.4 b 12.9 a -
200 0 16.2 b - - - 1.88 b 52.6 c 13.2 a -
Year 2014
Control 16.8 60.5 22.8 2.01 a 52.2 a 11.4 a 0.95 a
50 5 - 4.7 56.7 38.6 2.38 b 67.6 ab 11.8 a 0.92 a
100 5 - 2.8 51.8 45.4 2.50 b 68.6 ab 11.8 a 0.79 a
200 5 - 6.5 43.7 49.7 2.48 b 82.1 b 11.3 a 0.78 a
1Days after 50% blooming in 2010 and days after 80% blooming in 2014.
2Within each column, different letters indicate significant differences with Tukey-Kramer multiple comparison test (P≤0.05).
In 2014, the application of 100 mg L-1 GA was more effective in increasing fruit size
and fruit weight in comparison with 50 L-1 GA. However, the application of 200 mg L-1 GA
resulted in similar fruit size and weight as of 100 mg L-1 GA in both 2010 and 2014.
Therefore, we consider that the most suitable GA application rate in increasing fruit size of
highbush blueberry cultivar ‘Spartan’ is approximately 100 mg L-1.
The results of the experiment on timing of GA application are shown in Table 2. In
2010, fruit diameter and fruit weight were highest when GA was applied at 5 DAB. In 2013,
the proportion of large fruit (>18 mm) was highest following GA application at 5 DAB,
although there was no significant difference in fruit weight among the 0, 5, and 10 DAB
applications. Hence, we conclude that 5 DAB is the most effective stage at which to apply GA
to increase fruit size in highbush blueberry ‘Spartan’.

Table 2. Response of highbush blueberry cultivar ‘Spartan’ to GA applied at various timings.

Fruit size Soluble
GA application Fruit Fruit Titratable
distribution (%) Number of solid
diameter weight acid
Timing Rate <15 15-18 >18 seeds fruit-1 content
(mm) (g) (%)
(DAB1) (mg L-1) (mm) (mm) (mm) (°Brix)
Year 2010
Control 13.9 a2 - - - 1.25 a 23.1 a 12.8 ab -
0 200 15.8 b - - - 1.83 b 52.0 b 13.3 b -
5 200 16.9 c - - - 2.25 c 55.0 b 12.1 a -
10 200 15.6 b - - - 1.82 b 55.4 b 12.8 ab -
20 200 15.1 ab - - - 1.57 ab 45.7 b 12.7 ab -
Year 2013
Control - 59.4 34.4 6.3 1.36 a 2.3 a 12.9 a 0.75 a
0 100 - 32.7 62.7 4.5 1.76 ab 11.5 ab 11.7 a 0.67 a
5 100 - 24.0 64.5 11.6 1.92 b 13.5 ab 12.3 a 0.75 a
10 100 - 32.0 64.1 3.9 1.83 b 20.1 b 11.7 a 0.72 a
1Days after 50% blooming in 2010 and days after 80% blooming in 2013.
2Within each column, different letters indicate significant differences with Tukey-Kramer multiple comparison test (P≤0.05).

  185
CPPU application
CPPU applications of 10 or 20 mg L-1 significantly increased fruit size and weight in the
two treatment years (2010 and 2014) compared to the untreated control (Table 3). In 2014,
application of 5 mg L-1 CPPU was less effective for increasing fruit size and weight. Previous
studies reported that application of 5 mg L-1 CPPU to various blueberry cultivars was less
effective at increasing fruit size compared to higher CPPU rates (Williamson and NeSmith,
2007; Fujisawa et al., 2011; Retamales et al., 2014). Therefore, we suggest that the effective
CPPU application rate for increasing fruit size in highbush blueberry ‘Spartan’ is at least 10
mg L-1. However, we observed malformed leaves that had wavy leaf margins and a convex
surface between veins, following CPPU application at 10 and 20 mg L-1 (Figure 1).

Table 3. Response of highbush blueberry cultivar ‘Spartan’ to CPPU applied at various

concentrations.
Fruit size
CPPU application Fruit Fruit Soluble Titratable
distribution (%) Number of
diameter weight solid content acid
Rate Timing <15 15-18 >18 seeds fruit-1
(mm) (g) (°Brix) (%)
(mg L-1) (DAB1) (mm) (mm) (mm)
Year 2010
Control 14.1 a2 - - - 1.33 a 20.8 a 13.1 a -
10 10 16.5 b - - - 2.07 b 35.6 b 12.4 b -
20 10 17.6 c - - - 2.33 b 36.9 b 11.5 c -
Year 2014
Control 16.8 60.5 22.8 2.01 a 52.2 a 11.4 a 0.95 a
5 10 - 12.4 55.8 31.8 2.19 a 51.3 a 11.4 a 0.93 a
10 10 - 5.6 46.4 48.1 2.52 b 48.6 a 11.6 a 1.02 a
20 10 - 4.1 40.8 55.1 2.72 c 55.4 a 11.9 a 0.98 a
1Days after 50% blooming in 2010 and days after 80% blooming in 2014.
2Within each column, different letters indicate significant differences with Tukey-Kramer multiple comparison test (P≤0.05).

Figure 1. Normal leaf (left) and malformed leaf (right). Malformed leaves were frequently
observed on shoots treated with 10 or 20 mg L-1 CPPU.

The number of seeds per fruit increased significantly compared to the control after
CPPU application in 2010. Soluble solid contents were altered significantly by the treatment
in 2010 but not in 2014; titratable acid was not significantly different in 2014.
With regard to the timing of CPPU application, the results in 2010 demonstrated that
CPPU application at 10 DAB was the most effective for increasing fruit diameter and weight
compared to other timings (Table 4). A previous study on rabbiteye blueberry, in which
various application timings from 0 to 28 DAB were tested, demonstrated that later timings
led to higher fruit weights (NeSmith, 2008). Nevertheless, highbush blueberry ‘Spartan’
showed little response to CPPU applications at 20 DAB in our experiment. In 2014, the fruit
harvested from shoots treated at 10 DAB had the lowest proportion of small fruit (<15 mm)
and had the highest fruit weight among all the tested treatments. These results indicated

186
that the optimum CPPU application timing for increasing fruit size in the ‘Spartan’ cultivar is
approximately 10 DAB.

Table 4. Response of highbush blueberry cultivar ‘Spartan’ to CPPU applied at various

timings.
Fruit size
CPPU application Fruit Fruit Soluble solid Titratable
distribution (%) Number of
diameter weight content acid
Timing Rate <15 15-18 >18 seeds fruit-1
(mm) (g) (°Brix) (%)
(DAB1) (mg L-1) (mm) (mm) (mm)
Year 2010
Control 13.9 a2 - - - 1.25 a 23.1 a 12.8 a -
0 20 17.6 b - - - 2.42 bc 39.8 b 12.2 a -
5 20 17.1 b - - - 2.26 b 37.2 b 12.4 a -
10 20 17.9 b - - - 2.66 c 41.7 b 12.2 a -
20 20 14.7 a - - - 1.52 a 21.9 a 12.7 a -
Year 2013
Control - 59.4 34.4 6.3 1.36 a 2.3 a 12.9 a 0.75 a
0 10 - 67.9 27.4 4.7 1.49 a 5.1 ab 12.3 a 0.65 a
5 10 - 49.1 41.1 9.8 1.48 a 15.3 b 12.0 a 0.61 a
10 10 - 34.6 63.2 2.3 1.68 a 4.9 ab 11.6 a 0.70 a
1Days after 50% blooming in 2010 and days after 80% blooming in 2013.
2Within each column, different letters indicate significant differences with Tukey-Kramer multiple comparison test (P≤0.05).

Combined application of GA and CPPU

Blueberry fruit harvested from shoots given the combined GA and CPPU treatment had
a very high rate of large fruit (>18 mm) and also had the highest fruit weight among the
treatments in 2014 (Table 5). In 2016, fruit diameter, fruit weight, and number of seeds per
fruit were highest for the combined application compared to the control and to single
applications of GA and CPPU. Therefore, we conclude that the combined application of GA
and CPPU was very effective at increasing fruit size in the highbush blueberry ‘Spartan’.

Table 5. Response of highbush blueberry cultivar ‘Spartan’ to the combined application of

GA and CPPU.
Fruit size
Treatment Fruit Fruit Soluble solid Titratable
distribution (%) Number of
(rate (mg L-1), diameter weight content acid
<15 15-18 >18 seeds fruit-1
timing (DAB1)) (mm) (g) (°Brix) (%)
(mm) (mm) (mm)
Year 2014
Control - 11.5 60.7 27.9 2.09 a 47.2 a 11.4 a 0.88 a
GA (100, 5) - 4.8 50.4 44.8 2.56 b 62.8 a 12.3 a 0.81 a
CPPU (10, 10) - 5.6 31.8 62.6 2.90 c 57.1 a 12.4 a 0.83 a
GA + CPPU - 1.3 20.8 77.9 3.12 c 61.3 a 12.6 a 0.85 a
Year 2016
Control 16.4 a2 - - - 2.02 a 14.0 a 11.4 a 0.49 a
GA (100, 5) 17.6 b - - - 2.49 b 27.6 b 11.0 a 0.53 a
CPPU (20, 10) 17.9 b - - - 2.55 b 22.0 ab 10.9 a 0.59 a
GA + CPPU 18.4 c - - - 2.82 c 39.7 c 11.1 a 0.63 a
1Days after 80% blooming.
2Within each column, different letters indicate significant differences with Tukey-Kramer multiple comparison test (P≤0.05).
On the other hand, no significant differences were observed for soluble solid contents
or titratable acids in either year. Hence, we suggest that application of GA and CPPU, whether
as a combined treatment or as single applications, had no effect on internal fruit quality of
the ‘Spartan’ cultivar.
The combined application of GA and CPPU at the optimal rate and timing increased

  187
fruit weight in ‘Spartan’ by 40-49% compared to untreated controls. The combined
application of GA and CPPU might therefore improve the marketability of blueberry fruit.

CONCLUSIONS
The following conclusions can be drawn from the study:
- Both GA and CPPU applications increase fruit size, fruit weight, and number of seeds
per fruit in highbush blueberry cultivar ‘Spartan’.
- From the point of view of increasing fruit size, the most suitable GA application is
approximately 100 mg L-1, and the most effective time for GA application is
approximately 5 DAB.
- Similarly, the most effective CPPU application rate is at least 10 mg L-1, and the best
time for CPPU application is approximately 10 DAB.
- A combined application of 100 mg L-1 GA at 5 DAB and 10 mg L-1 or 20 mg L-1 CPPU
at 10 DAB is very effective for increasing fruit size without any impact on soluble
solid content and acidity of the fruit.

Literature cited
Cano-Medrano, R., and Darnell, R.L. (1998). Effect of GA3 and pollination on fruit set and development in
rabbiteye blueberry. HortScience 33 (4), 632–635.
Fujisawa, H., Kawai, Y., and Ishikawa, K. (2011). Effects of GA and CPPU applications on fruit set and fruit
characteristics of northern highbush blueberries. J. Agric. Sci. Tokyo Univ. Agric. 56 (2), 118–124.
Mainland, C.M., and Eck, P. (1969). Fruiting response of the highbush blueberry to gibberellic acid under field
conditions. J. Am. Soc. Hortic. Sci. 94, 21–23.
Merino, R., Serri, H., and Holzapfel, J. (2002). Effect of GA3 and CPPU on fruit of ‘Tifblue’ rabbiteye blueberry
(Vaccinium ashei R.). Acta Hortic. 574, 239–243 https://doi.org/10.17660/ActaHortic.2002.574.36.
NeSmith, D.S. (2002). Response of rabbiteye blueberry (Vaccinium ashei Reade) to the growth regulators CPPU
and gibberellic acid. HortScience 37 (4), 666–668.
NeSmith, D.S. (2005). Use of plant growth regulators in blueberry production in the southeastern U.S.: A review.
Int. J. Fruit Sci. 5 (3), 41–54 https://doi.org/10.1300/J492v05n03_05.
NeSmith, D.S. (2008). Effects of timing of CPPU applications on rabbiteye blueberries. HortScience 43 (5), 1446–
1448.
NeSmith, D.S., and Adair, H.M. (2004). Rabbiteye blueberry field trials with the growth regulator CPPU. Small
Fruit Rev. 3 (1-2), 183–191 https://doi.org/10.1300/J301v03n01_18.
NeSmith, D.S., Krewer, G., Rieger, M., and Mullinix, B. (1995). Gibberellic acid-induced fruit set of rabbiteye
blueberry following freeze and physical injury. HortScience 30 (6), 1241–1243.
Retamales, J.B., Lobos, G.A., Romero, S., Godoy, R., and Moggia, C. (2014). Repeated applications of CPPU on
highbush blueberry cv. Duke increase yield and enhance fruit quality at harvest and during postharvest. Chil. J.
Agric. Res. 74 (2), 157–161 https://doi.org/10.4067/S0718-58392014000200005.
Serri, H., and Hepp, R. (2006). Effect of the growth regulator CPPU on fruit quality and fruit ripening of highbush
blueberries. Acta Hortic. 715, 279–283 https://doi.org/10.17660/ActaHortic.2006.715.40.
Williamson, J.G., and NeSmith, D.S. (2007). Effects of CPPU applications on southern highbush blueberries.
HortScience 42 (7), 1612–1615.

188