Growth, bolting and yield of garlic (Allium sativum L.) in response to clove chilling treatment

Scientia Horticulturae 194 (2015) 43–52

Contents lists available at ScienceDirect

Scientia Horticulturae journal homepage: www.elsevier.com/locate/scihorti

Growth, bolting and yield of garlic (Allium sativum L.) in response to clove chilling treatment Cuinan Wu, Mengyi Wang, Yinxin Dong, Zhihui Cheng ∗ , Huanwen Meng College of Horticulture, Northwest A&F University, Yangling, Shaanxi 712100, China

a r t i c l e

i n f o

Article history: Received 19 March 2015 Received in revised form 7 July 2015 Accepted 13 July 2015 Keywords: Bolting Garlic growth Clove Vernalization Yield

a b s t r a c t Garlic (Allium sativum L.) has been long favored as a seasoning and as a seasonal vegetable all over the world. However, the production of fresh garlic scapes and bulbs is extremely limited by the climate and region, especially the temperature and photoperiod, under natural production conditions, and the uncertainty of the vernalization characteristic of garlic has been a key restriction to the developmental regulation of bolting and bulbing. A thorough understanding of the vernalization conditions for garlic growth should improve our knowledge of the bolting and bulbing processes. This research was conducted to assess the response of the clove to chilling treatment and further demonstrate whether chilling treatment on garlic clove could substitute the vernalization requirements for garlic plant. Before planting, the cloves (cv. G064) were subjected to low temperature (5 ◦ C, 10 ◦ C, 15 ◦ C) for different durations (20 days, 40 days or 60 days) for chilling treatment. The control cloves or plants remained in the 20–22 ◦ C regime throughout. Then, the plant growth, bolting and garlic yield were evaluated during field growing. The results indicate that pre-plant chilling treatment significantly affects garlic plant growth and development, and could act as the vernalization and enable the garlic plant to bolt without undergoing chilling during plant growth. Lower temperatures and longer treatment durations significantly enhanced the leaf elongation, shortened the growth period (5 ◦ C or 60 days), promoted the rate of one-clove bulbs (5 ◦ C or 40 days), stimulated bolting and increased the bolting rate (10 ◦ C) compared with the control. However, the bolting rate and low-temperature treatment did not follow a simple linear relation, as lower temperatures (5 ◦ C) or higher temperatures (15 ◦ C) were not beneficial to garlic bolting. It was found that garlic plants subjected to a treatment of 10 ◦ C for 20/40 days or 5 ◦ C for 20 days had comparatively higher bolting rates. Nevertheless, the garlic bulb yield and mean bulb weight decreased significantly as a function of the increasing period of the chilling treatment compared with the control. The treatment of 5 ◦ C for 20 days had the greatest yield among all the treatments, second only to the control. These findings provide strong evidence for the potential cultivation of garlic of various cultivars in various climates or seasons, making off-season cultivation possible. Other researchers could use a similar methodology to shorten the period needed until harvest or even produce desirable scapes and bulbs in the warm season. © 2015 Elsevier B.V. All rights reserved.

1. Introduction Garlic (Allium sativum L.) is a well-known spice and seasonal vegetable with antiseptic and antibacterial properties (Sendl, 1995). In recent years, there has been a significant increase in the productivity and quality of garlic, with the garlic cultivation area exceeding 804 thousand hectares with over 230 thousand tons yield (FAO, 2010, see Supplementary material 1) in China. However, the production of garlic scapes and bulbs is extremely limited

∗ Corresponding author. Fax: +86 29 87082613. E-mail address: [email protected] (Z. Cheng). http://dx.doi.org/10.1016/j.scienta.2015.07.018 0304-4238/© 2015 Elsevier B.V. All rights reserved.

by the climate and region, especially the temperature and photoperiod, under natural production conditions. Moreover, little is known about the developmental mechanism of the garlic flower bud and stalk, especially the response of garlic to vernalization treatment, which restricts the development of scape and bulb production in various climates. Thus, a thorough understanding of the vernalization conditions for garlic growth should improve our knowledge of the bolting process and facilitate the production of fresh scapes and bulbs in various climates. Temperature and photoperiod are important environmental conditions for good development of the crop (Filgueira, 2008). Low temperature is the main factor for garlic bolting that affects bulb growth, resulting in increases or delays in development (Mojtahedi

44

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

et al., 2013). Vernalization, assumed to play an important role in plant bolting, is the process by which floral induction in plants is promoted due to the plants being exposed to chilling for a certain period of time (Ritz et al., 2010). This process is categorized as either seed vernalization (in which low temperatures are sensed by germinating seeds, such as in Triticum aestivum, Brassica pekinensis, Spinacia oleracea and Raphanus sativus) or green plant vernalization (in which low temperatures are sensed by plants at a certain age with green leaves, such as in garlic, Allium cepa, Allium fistulosum, Apium graveolens and Brassica oleracea) (Song et al., 2010; Dong et al., 2013). Trevaskis et al. (2007) reported that several weeks of cold exposure were often sufficient to promote flowering, but longer durations could accelerate flowering to a greater degree, up to the point when the vernalization response became saturated. It was reported that the optimum conditions to fully vernalize onion cultivars were under low-temperature (–11 ◦ C) treatments for 7–90 days (Khokhar et al., 2007). The role of lowtemperature storage for growth and development in other bulbs has also been well demonstrated (Mojtahedi et al., 2012, 2013). Tulip (Tulipa gesneriana) bulbs require a low-temperature treatment (4–5 ◦ C) for shoot elongation and flower formation (Le Nard and De Hertogh, 1993). Low-temperature exposure also influences specific phases of growth, and the optimum temperature and exposure time diverges with the growth stage (Lucidos et al., 2014). In Allium, flower scape development reaches a maximum at 11 ◦ C, whereas in the tulip flower scape, maximum elongation occurs at 0–9 ◦ C for 10–12 weeks (Le Nard and De Hertogh, 1993). However, the cold requirement of bulbs for garlic bolting, bulbing or flowering has not been thoroughly researched recently. Garlic genotypes are categorized as non-bolting, semi-bolting, and bolting types (Takagi, 1990; Kamenetsky and Rabinowitch, 2001; Etoh and Simon, 2002; Kamenetsky et al., 2004a,b). Bolting in garlic occurs only in plants that experience the appropriate environmental conditions (Takagi, 1990). The bolting and bulbing of garlic are influenced by the day length and the temperature to which the dormant cloves or growing plants are exposed before bolting begins (Bandara et al., 2000). Nakamura (1985) and Takagi (1990) proposed that a minimum critical cold period was needed for floral induction in garlic. In general, low initial temperatures followed by long days are essential for bolting, scape elongation and the formation of bulbs and cloves (Takagi, 1990). Lee et al. (2007, 2008) reported that treating Asiatic hybrid lily (Lilium x elegans) bulbs with an optimum vernalization temperature of 5 ◦ C increased the number of flower buds and also accelerated flowering, while temperatures higher than 12.5 or 15 ◦ C during bulb vernalization treatments reduced the number of flower buds. The vernalization requirement of garlic is generally met in China by overwintering in the field for fall-planted cloves or by experiencing the cool early spring for spring-planted cloves. As a nutrient storage organ, the clove could be considered a micro garlic plant. However, whether the clove can sense low temperatures and saturate the vernalization is still unknown. The research of Bandara et al. (2000) showed that better cloving and higher bulb yields were obtained if garlic cloves were treated at 4 ◦ C for 45 or 60 days prior to field or greenhouse planting, respectively. Pyo et al. (1979) indicated that the most striking effect of vernalization in garlic was the increase in earliness, especially in cultivars with a greater low-temperature requirement for development (Motaz et al., 1971). Siddique and Rabbani (1985) reported that the treatment of garlic cloves at 6 ◦ C for 50 days before planting increased the bulb size and yield. Nevertheless, much lower temperatures tend to initiate the secondary growth of garlic, which is defined as the lateral bud differentiation into secondary plants on the axils of the outer layer leaves of the primary plant or the continuous growth of the primary plant’s clove or bulbil buds into secondary plants instead of turning dormant at the end of the sea-

son (Cheng et al., 1991). The secondary growth of the garlic plant significantly affects the commercial performance of fresh scapes and bulbs. Taken together, the low-temperature treatment of cloves prior to planting significantly affects garlic bulbing, cloving and yield (Bandara et al., 2000). However, the relationship between the chilling treatment of the clove and vernalization has not yet been elucidated. Moreover, whether the cold accumulation of the chilling treatment is enough for garlic vernalization and bolting of the clove needs to be researched. The objective of this research was to assess the influence of different pre-planting chilling treatments (by temperature and duration) of a garlic clove on growth, bolting rate and bulb yield and then further demonstrate whether chilling treatment on garlic clove could substitute the vernalization requirements for garlic plant. 2. Materials and methods 2.1. Experimental site description The field experiment was carried out under a plastic tunnel at the Horticultural Experimental Station (34◦ 16 N, 108◦ 4 E) of Northwest A&F University, Yangling, Shaanxi Province, China in 2011. The climate of the region is classified as a subtropical monsoon climate with hot summers and cool winters (Fig. 1). The annual average temperature is 12.9 ◦ C and the frost-free period is over 200 days. The chemical characteristics of the soil were as follows: pH (1:1 water) 7.83, electrical conductivity (1:5 soil/water) 239.1 ␮S cm−1 , available nitrogen (N) 56.32 mg kg−1 , available phosphorus (P) 52.57 mg kg−1 and available potassium (K) 224.90 mg kg−1 . 2.2. Experimental design The main local cultivated cultivar G064 (bolting type) was chosen as the material. The phenotype of cv. G064 was as follows: plant standing height 70–80 cm, leaf number 13–15, bulb weight 50 g, bulb diameter 5 cm, 12–13 cloves arranged in two whorls (outer whorl 6–7 cloves, inner whorl 5–7 cloves), bolting rate 80–85%, growth period 250 days in overwinter planting and dormancy duration 40 days (see Supplementary material 2). Normally, the cultivar needs 30–40 days under 0–4 ◦ C or 50–60 days under 10 ◦ C at the four-leaf age for vernalization (Song et al., 2010). After that period, a long photoperiod (≥13 h) and higher temperature (20 ◦ C) are needed for the bolting and bulbing of garlic. The cloves used for experiment were planted in the field mentioned above in September 2010 and harvested in early May 2011. Then, the bulbs were subjected to the cure process in a wellventilated room at room temperature (22 ± 2 ◦ C) for approximately 2 months. Uniform sound cloves were selected and sterilized using 6% Dacotech (75% chlorothalonil, Syngenta, China) for a 20-day interval and then placed in growth chambers (Ningbo Jiangnan Instrument Factory, Zhejiang Province, China) at 5 ◦ C (T5 ), 10 ◦ C (T10 ) or 15 ◦ C (T15 ) and 80% RH for chilling treatment. The chilling duration at each temperature was 20 days (D20 ), 40 days (D40 ) or 60 days (D60 ). After the chilling treatment, the cloves were immediately planted in the field on September 21, 2011 at 5 cm depth, with the plant spacing 5 cm and row spacing 20 cm under a plastic tunnel using a random complete block design with three replications. Another sample of the cloves stored continuously at ambient temperature (22 ± 2 ◦ C) and an RH of 70% was also planted to serve as the untreated control (Tck Dck ). The plot area was defined by a 1.5 m wide and 3.5 m long bed, with six planting rows. Each replication per treatment had two rows in one plot with 80 cloves. As the garlic

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

45

Fig. 1. Air temperature in the field in Yangling, Shaanxi Province, China from 2011 to 2012. The data shows the mean of the highest and lowest temperatures each day within a month.

plant grew, the temperature dropped, a lower plastic tunnel was built, and an electrical heating wire with a controller was laid to maintain the temperature at 22 ± 2 ◦ C/16 ± 2 ◦ C (day/night). Before planting, a base fertilizer of 1.5 kg “Pengdixin” (organic matter ≥ 30%, N + P2 O + K2 O ≥ 4%, humic acid ≥ 20%, organic sylvite ≥ 5%, Zhengzhou, Henan Province, China), 0.25 kg Stanley (N:P:K = 21:10:11, Yimeng, Shandong Province, China) and 0.4 kg ammonium hydrogen carbonate (Hanzhong, Shaanxi Province, China) were applied by broadcasting, and then the soil was deeply ploughed and uniformly tilled. Calcium ammonium nitrate potassium (Xuzhou, Jiangsu Province, China) was used as topdressing and applied 5 times during the growth period of the garlic at 0.125 kg per plot. Water was supplemented by irrigation regularly throughout the growing season and stopped five days before the harvest. Plant health was controlled with products based on Maneb and Thiometon-based insecticides. Weeds were manually removed. The standard agronomic practices were equally performed on all the experimental plots. Samples of 15 plants of each block were taken for the recording of garlic plant standing height, leaf sheath length and leaf sheath diameter at 103, 136, 146 and 160 days after planting. In the following May, the garlic bolting rate, garlic bulb yield and garlic bulb character were determined.

harvesting a 2 m long section in the center of the middle row of each plot (Bandara et al., 2000). 2.5. Assessment of garlic bolting rate, rate of one-clove bulb and growth period The bolting rate was calculated by the ratio of the number of harvested scapes to the total number of plants in one experimental unit. The harvest date was determined by the yellowing and partial drying of the canopy and/or plant falling. Then, the days of the

2.3. Measurement of garlic plant standing height, leaf sheath length and leaf sheath diameter The garlic plant standing height, leaf sheath length and leaf sheath diameter were evaluated in the laboratory using a measuring tape (0.01 cm) or electronic vernier caliper (0.01 mm, for leaf sheath diameter), and the number of plant leaves was counted by visual observation. 15 Plants of each block were taken with three replications. Garlic plant standing height was measured from the basal part of the leaf sheath (i.e. also the top of the bulb) to the tip of the longest leaf. Leaf sheath length was recorded from the basal part of leaf sheath to the beginning of the first leaf. The largest diameter of the leaf sheath (near 1 cm above the top of the bulb) was recorded as leaf sheath diameter (see Fig. 2). 2.4. Evaluation of garlic bulb character and yield The mean bulb weight was recorded using 20 randomly selected bulbs from each experimental unit. Bulb yields were determined by

Fig. 2. Measuring plant height, leaf sheath length and leaf sheath diameter.

46

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

growth period were calculated from date of planting in the field to the harvest date. After harvesting, the plants were subjected to a cure process at room temperature (22 ± 2 ◦ C) for approximately 2 weeks, and the total yield in ha−1 and the percentage of one-clove bulbs were then recorded (Resende et al., 2011). 2.6. Statistical analysis The data were subjected to analysis of variance (ANOVA) as a 4 × 4 (temperature × its duration) factorial structure, using the SPSS 17.0 software package. Mean separations among treatments were performed by the Fisher’s protected least significant difference (LSD) multiple range tests at 5% probability. 3. Results 3.1. Main effect of low temperature and its duration treatment on plant standing height, leaf sheath length and leaf sheath diameter The main effect of pre-planting low temperature and its duration treatment of garlic clove on plant standing height, leaf sheath length and leaf sheath diameter were analyzed (Table 1). All the studied factors, including temperature and temperature duration significantly influenced the garlic plant standing height, leaf sheath length and leaf sheath diameter. Meanwhile, the level of significant effect varied among different sampling dates. As garlic plant grew till 146 days after planting, the significant effect of low temperature treatment on plant standing height became higher (P < 0.05 for 103 days, P < 0.01 for 136 days, P < 0.001 for 146 days); while no significant effect was recorded when sampled 160 days after planting (P > 0.05). Main effect of chilling duration on plant standing height was also highly significant reaching 0.001 level, except for plants sampled 160 days after planting (P < 0.01). Besides, the test temperature or its duration treatment significantly affected plant leaf sheath length reaching 0.001 level of all three sampling dates. However low temperature treatment only significantly affected leaf sheath diameter when sampled 103 days after planting (P < 0.001); no significant influenced was obtained in the rest sampling dates. The significant effect of chilling duration treatment on leaf sheath diameter showed the similar pattern: significant effect was only shown when sampled 103 (P < 0.01) and 136 days after planting (P < 0.05). Moreover the lower temperature or longer duration presented stronger enhancing effects on plant standing height, and leaf sheath length. T5/10 or D60 showed significantly greater plant standing height and leaf sheath length; while T15 or D20 showed no significant difference from CK in all the sampling dates. 3.2. Interaction effects of low temperature and its duration treatment on plant standing height, leaf sheath length and leaf sheath diameter The interactions of temperature × duration (T × D) had significant effect on plant standing height when sampled 103 and 146 days after planting (P < 0.05); while no significant effect was recorded when sampled 136 or 160 days after planting (Table 2). Meanwhile the combination treatment of T × D had a highly significant influence on plant leaf sheath length except for plants sampled 169 days after planting (Table 3). While significant effect of T × D on leaf sheath diameter was only presented when sampled 103 days after planting (P < 0.01); no significant effects was recorded in the rest sampling dates (Table 4).

As shown in Tables 2 and 3, plant standing height and leaf sheath length was significantly promoted with the decrease in the treated temperature and increase in the low temperature duration compared to the control. T5 D60 and T10 D60 most effectively influenced the plant standing height and leaf sheath length (Tables 2 and 3). Likewise, the pre-planting clove chilling treatment significantly improved the leaf sheath diameter, especially for T5 D40 and T5 D60 , compared with the control when sampled 103 days after planting (Table 4). The change trend was the same for the plant standing height; the lower temperature (T5 ) and longer duration (D40/60 ) had a strong enhancing effect on garlic leaf sheath diameter. However, no significant differences in leaf number were found among the treatments (data not shown). It was observed that at the early growth stage (from emergence till 170 days after planting), the plant grew well as a result of the lower-temperature and longer-duration treatments (T5 D40 , T5 D60 and T10 D60 ). However, at the late growth stage (from 180 days after planting till harvest), the plant growth of T15 or D20 excelled, while the plants of T5 D40 , T5 D60 and T10 D60 had significant occurrences of secondary growth and premature senescence (data not shown). These results suggest that lower-temperature and longerduration treatments of the cloves significantly increased plant elongation growth while having no effect on the canopy growth (leaf number) of the plant. 3.3. Main effect of low temperature and its duration treatment on bolting rate, growth period, rate of one-clove bulb and yield of garlic plants Table 5 presented main effect of pre-planting low temperature and its duration treatment of garlic clove on garlic bolting rate, growth period, rate of one-clove bulb and yield. Highly significant influence was obtained in all the studying factors (temperature, duration; P < 0.001). It was obvious that low temperature or its duration treatment significantly increased the bolting rate and decreased the days needed till harvest compared with control. T10 or D20 had the highest bolting rate, followed by T15 or D40. The growth period dropped with the decrease of test temperature or increase of temperature duration (T5 < T10 < T15 , D60 < D40 < D20 ); T15 showed no significant difference from the control. Oppositely the number of one-clove bulb increased as the test temperature went lower and the duration became longer (T5 > T10 > T15 , D40 > D60 > D20 ); control was recorded with the lowest one-clove bulb rate (0). After clove chilling treatment, bulb yield was significantly reduced compared with control. The lower test temperature or longest duration showed the strongest inhibiting effect on bulb yield (T10 < T5 < T15 < CK, D60 < D40 < D20 < CK). A similar change pattern was obtained when analyze the significant effect of test temperature or duration on mean bulb weight. 3.4. Interaction effects of low temperature and its duration treatment on bolting rate and growth period of garlic plants The interaction effects of T × D on garlic bolting rate and growth period were highly significant, reaching 0.001 level (Table 6). It was recorded that all the treatments increased the bolting rate compared with the control (3.3%). This result suggests that a low-temperature treatment on the clove could act as the vernalization and enable the garlic plant to bolt without undergoing chilling during plant growth. The bolting rate and low-temperature treatment does not follow a simple linear relation. As shown in Table 6, T5 D40 and T5 D60 with the lowest test temperature and relatively longer temperature duration presented the lowest bolting rate compared with the other chilling treatments. The higher secondary growth rate and premature senescence might account for this phenomenon (data

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

47

Table 1 Main effect of low temperature and its duration treatment on plant standing height, leaf sheath length and leaf sheath diameter sampled 103, 136, 146 or 160 days after planting. Treatment

Plant standing height (cm)

Plant leaf sheath length (cm)

Plant leaf sheath diameter (mm)

103

136

146

160

103

136

146

160

103

136

146

160

Grand mean

64.1

68.0

67.8

71.9

11.2

14.1

15.5

16

8.8

9.9

10.9

9.1

Temperature (◦ C) CK 5 10 15

59.9 ab 67.6 a 67.3 a 58.9 b

62.1 bc 75.9 a 68.6 b 61.6 c

59.8 c 76.9 a 68.1 b 61.0 bc

59.3 b 75.3 a 78.6 a 66.0 ab

9.4 b 12.5 a 12.3 a 9.5 b

10.9 c 17.0 a 15.0 b 11.4 c

12.1 c 19.3 a 16.4 b 12.0 c

12.3 b 18.9 a 17.7 a 12.6 b

9.0 b 9.7 a 8.4 bc 8.1 c

10.1 a 10.2 a 9.8 a 9.8 a

9.9 a 14.2 a 9.4 a 9.5 a

8.5 a 8.9 a 9.0 a 9.7 a

Duration (day) CK 20 40 60

59.9 b 57.7 b 62.3 b 73.9 a

62.1 b 60.5 b 66.0 b 79.6 a

59.8 c 58.0 c 69.8 b 78.2 a

59.3 c 62.8 bc 74.7 ab 82.5 a

9.4 b 9.6 b 11.0 b 13.6 a

10.9 c 12.1 c 14.2 b 16.9 a

12.1 c 12.0 c 15.8 b 20.0 a

12.3 c 13.2 bc 16.4 b 19.7 a

9.0 a 8.3 b 8.7 ab 9.2 a

10.1 ab 9.3 b 9.7 ab 10.7 a

9.9 a 9.1 a 11.0 a 13.0 a

8.5 a 8.6 a 9.4 a 9.6 a

*

**

***

NS

***

***

***

***

***

NS

***

***

***

**

***

***

***

***

**

*

NS NS

NS NS

F-Test Temperature (T) Duration (D)

NS stands for no significant difference at 0.05 level. CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). Different letters indicate significant differences between means within columns at the 5% probability level by LSD. * Significant at 0.05 level (ANOVA and LSD’s multiple range test). ** Significant at 0.01 level. *** Significant at 0.001 level.

not shown). Garlic plants under T10 or D20/40 showed relatively higher bolting rates (except T5 D40 ). T10 D40 recorded the highest bolting rate (66.7%), followed by T5 D20 (59.3%) and T10 D20 (59.3%). Clearly, lower-temperature and longer-duration treatments significantly shortened the growth period (days from planting to harvest) of garlic (Table 6). However, T10 D20 and T15 exhibited no significant difference from the control. T5 D60 had the shortest period for garlic growth (199 days), and T5 D40 was the second shortest (209 days). It seems that the clove chilling treatment accelerated the maturation of the garlic plant. 3.5. Interaction effects of low temperature and its duration treatment on rate of one-clove bulb and yield of garlic plants Highly significant influences were also obtained when analyzed the interaction effect of T × D on rate of one-clove bulb and yield of garlic plants (P < 0.001) Table 6 demonstrated that pre-planting chilling treatment on the clove gave rise to a significantly greater number of one-clove

bulbs (not desirable in production due to their small size) than the control (0). T5 D40 and T5 D60 recorded the largest number of one-clove bulbs, 17 and 11, respectively. The results indicate that the control treatment, not subjected to low temperature [stored continuously at ambient temperature (22 ± 2 ◦ C) and grew at 22 ± 2 ◦ C/16 ± 2 ◦ C (day/night) throughout], gave rise to a normal bulb (bulb with multiple cloves). This contradicts the traditional view of chilling (lower than 20 ◦ C) as being essential for cloving and bulbing of garlic. Furthermore, the bulb yield decreased after the pre-planting clove chilling treatment (Table 6). The control presented the most productive performance. Meanwhile, the lower temperature or a longer duration of low-temperature treatment presented a lower bulb yield; plants under T15 or D20 were preponderant in bulb yield (second to the control) and T5 D20 had the greatest yield among all the chilling treatments. These results indicate that lower-temperature and longer-duration treatments decrease the yield but increase the number of one-clove bulbs.

Table 2 Interaction effects of low temperature and its duration treatment on plant standing height sampled 103, 136, 146 or 160 days after planting. Clove chilling treatment ◦

Plant standing height (cm)

Temperature ( C)

Duration (day)

103 days

136 days

146 days

160 days

CK

CK

59.9 ± 6.4 cd

62.1 ± 2.3 cd

59.8 ± 7.2 cd

59.2 ± 5.5 c

5

20 40 60

54.8 ± 6.9 d 63.8 ± 3.3 bcd 84.1 ± 5.2 a

60.3 ± 8.2 d 73.3 ± 5.5 bc 94.2 ± 3.6 a

61.2 ± 1.1 cd 79.5 ± 0.3 b 90.1 ± 1.2 a

56.9 ± 10.9 c 79.1 ± 8.6 ab 89.9 ± 4.7 a

10

20 40 60

56.9 ± 4.6 cd 69.5 ± 1.6 bc 75.6 ± 3.9 ab

62.2 ± 1.4 cd 64.9 ± 4.7 cd 78.5 ± 2.7 b

54.8 ± 3.2 d 68.2 ± 2.4 c 81.2 ± 2.4 ab

67.9 ± 1.2 bc 77.5 ± 4.6 ab 90.3 ± 5.7 a

15

20 40 60

61.2 ± 4.0 cd 53.5 ± 0.7 d 62.0 ± 3.4 cd

58.9 ± 2.6 d 59.9 ± 2.5 d 66.1 ± 1.9 cd

58.1 ± 5.6 cd 61.6 ± 2.9 cd 63.2 ± 3.1 cd

63.5 ± 7.4 bc 67.3 ± 4.7 bc 67.4 ± 2.4 bc

*

NS

*

NS

F-Test Temperature × duration

The data are presented as the means ± standard error, and 15 plants of each block were taken with three replications. Different letters in the same column indicate significant differences at the 0.05 level (ANOVA and LSD’s multiple range test), n = 3. NS stands for no significant difference at 0.05 level. CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). * Significant at 0.05 level (ANOVA and LSD’s multiple range test).

48

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

Table 3 Interaction effects of low temperature and its duration treatment on plant leaf sheath length sampled 103, 136, 146 or 160 days after planting. Clove chilling treatment

Plant leaf sheath length (cm)

Temperature (◦ C)

Duration (day)

103 days

136 days

146 days

160 days

CK

CK

9.4 ± 1.0 e

10.9 ± 0.5 d

12.1 ± 0.5 e

12.3 ± 14 e

5

20 40 60

9.0 ± 0.7 e 12.0 ± 0.7 cd 16.4 ± 1.1 a

12.3 ± 0.6 cd 17.6 ± 0.7 b 21.1 ± 1.1 a

12.4 ± 0.2 e 19.0 ± 0.4 c 26.4 ± 0.5 a

13.4 ± 16 de 19.2 ± 12 bc 24.1 ± 37 a

10

20 40 60

10.0 ± 0.4 cde 12.3 ± 0.6 bc 14.5 ± 1.0 ab

13.1 ± 1.0 cd 14.2 ± 1.1 c 17.5 ± 1.5 b

11.6 ± 0.7 e 16.3 ± 0.8 d 21.4 ± 0.4 b

14.5 ± 0.3 de 17.3 ± 1.2 bcd 21.2 ± 0.7 ab

15

20 40 60

9.9 ± 1.2 de 8.9 ± 0.2 e 9.8 ± 0.5 de

11.0 ± 0.7 d 10.9 ± 0.2 d 12.2 ± 0.3 cd

11.9 ± 1.1 e 12.1 ± 0.6 e 12.1 ± 0.2 e

11.4 ± 1.2 e 12.5 ± 0.8 e 13.9 ± 0.4 de

**

**

***

NS

F-Test Temperature × duration

The data are presented as the means ± standard error, and 15 plants of each block were taken with three replications. Different letters in the same column indicate significant differences at the 0.05 level (ANOVA and LSD’s multiple range test), n = 3. NS stands for no significant difference at 0.05 level. CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). ** Significant at 0.01 level (ANOVA and LSD’s multiple range test). *** Significant at 0.001 level.

Table 4 Interaction effects of low temperature and its duration treatment on plant leaf sheath diameter sampled 103, 136, 146 or 160 days after planting. Clove chilling treatment

Plant leaf sheath diameter (mm)

Temperature (◦ C)

Duration (day)

103 days

136 days

146 days

160 days

CK

CK

9.0 ± 0.3 bc

10.1 ± 0.1 ab

9.9 ± 1.0 b

8.5 ± 0.3 ab

5

20 40 60

8.5 ± 0.3 cde 9.9 ± 0.2 ab 10.7 ± 0.4 a

9.5 ± 1.2 b 9.8 ± 0.5 ab 11.3 ± 1.0 a

9.5 ± 0.1 b 14.2 ± 4.2 ab 19.0 ± 7.9 a

8.0 ± 1.1 b 8.8 ± 1.2 ab 9.9 ± 0.5 ab

10

20 40 60

7.9 ± 0.4 e 8.5 ± 0.3 cde 8.9 ± 0.3 cd

9.0 ± 0.4 b 9.8 ± 0.1 ab 10.6 ± 0.2 ab

8.5 ± 0.3 b 9.3 ± 0.1 b 10.5 ± 0.4 b

8.6 ± 0.4 ab 9.0 ± 0.1 ab 9.5 ± 0.6 ab

15

20 40 60

8.5 ± 0.1 cde 7.6 ± 0.1 e 8.1 ± 0.4 de

9.4 ± 0.3 b 9.6 ± 0.3 ab 10.3 ± 0.6 ab

9.3 ± 0.6 b 9.6 ± 0.5 b 9.5 ± 0.4 b

9.3 ± 0.6 ab 10.2 ± 1.0 a 9.5 ± 0.5 ab

**

NS

NS

NS

F-Test Temperature × duration

The data are presented as the means ± standard error, and 15 plants of each block were taken with three replications. Different letters in the same column indicate significant differences at the 0.05 level (ANOVA and LSD ’s multiple range test), n = 3. NS stands for no significant difference at 0.05 level. CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). ** Significant at 0.01 level (ANOVA and LSD’s multiple range test).

Table 5 Main effect of low temperature and its duration treatment on bolting rate, growth period, rate of one-clove bulb, yield and mean bulb weight of garlic plants. Treatment

Bolting rate (%)

Growth period (day)

Rate of one-clove bulb (%)

Garlic bulb yield (t/ha)

Mean bulb weight (g)

Grand mean

46.1

225.6

9.6

5.6

36.5

Temperature ( C) CK 5 10 15

3.3 d 37.1 c 60.0 a 55.6 b

234.3 a 212.6 c 227.1 b 234.3 a

0d 22.5 a 6.6 b 3.1 c

7.9 a 5.0 c 4.9 d 6.0 b

49.7 a 34.9 c 33.6 d 36.5 b

Duration (day) CK 20 40 60

3.3 d 57.5 a 48.5 b 46.7 c

234.3 a 232.3 b 224.2 c 217.4 d

0d 6.8 c 13.8 a 11.5 b

7.9 a 6.6 b 5.7 c 3.7 d

49.7 a 42.1 b 34.8 c 28.2 d

***

***

***

***

***

***

***

***

***

***

◦

F-Test Temperature (T) Duration (D)

CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). Different letters indicate significant differences between means within columns at the 5% probability level by LSD. *** Significant at 0.001 level.

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

49

Table 6 Interaction effects of low temperature and its duration treatment on bolting rate, growth period, rate of one-clove bulb and yield of garlic plants. Clove chilling treatment

Bolting rate (%)

Growth period (day)

Rate of one-clove bulb (%)

Garlic bulb yield (t/ha)

CK

3.3 ± 0.7 h

234 a

0h

7.94 ± 0.12 a

5

20 40 60

59.3 ± 0.7 b 22.0 ± 1.2 g 30.0 ± 1.2 f

229 b 209 d 199 e

10.0 ± 0.2 c 34.2 ± 5.1 a 22.9 ± 2.8 b

7.32 ± 0.04 b 5.70 ± 0.03 f 2.15 ± 0.10 i

10

20 40 60

59.3 ± 1.3 b 66.7 ± 0.7 a 54.0 ± 1.2 e

234 a 229 b 219 c

5.3 ± 1.3 de 6.0 ± 1.8 e 8.4 ± 1.2 d

6.21 ± 0.10 c 5.55 ± 0.09 g 2.98 ± 0.08 h

15

20 40 60

54.0 ± 2.4 e 56.7 ± 0.7 c 56.0 ± 1.2 d

234 a 234 a 234 a

4.7 ± 1.8 f 1.4 ± 1.2 g 3.3 ± 1.4 f

6.21 ± 0.02 c 5.97 ± 0.12 d 5.95 ± 0.15 e

***

***

***

***

Temperature (◦ C)

Duration (day)

CK

F-Test Temperature × duration

The data are presented as the means ± standard error. Different letters in the same column indicate significant differences at the 0.05 level (ANOVA and LSD’s multiple range test), n = 3. CK stands for cloves stored continuously at ambient temperature (22±2 ◦ C). *** Significant at 0.001 level (ANOVA and LSD’s multiple range test). Table 7 Interaction effects of low temperature and its duration treatment on garlic bulb character. Clove chilling treatment

Garlic bulb character

Temperature (◦ C)

Duration (day)

Mean garlic bulb weight (g)

Garlic bulb diameter (mm)

Garlic bulb height (mm)

CK

CK

49.7 ± 5.8 a

48.9 ± 1.9 a

40.4 ± 0.8 a

5

20 40 60

44.1 ± 1.3 b 30.1 ± 5.5 cde 27.0 ± 5.5 de

48.4 ± 0.4 a 46.9 ± 2.6 b 45.6 ± 2.7 c

37.9 ± 1.1 b 32.8 ± 1.7 e 32.0 ± 1.4 f

10

20 40 60

40.0 ± 1.6 c 38.2 ± 0.5 cd 22.7 ± 0.6 e

45.4 ± 1.9 c 47.1 ± 0.5 b 41.7 ± 0.3 f

37.3 ± 0.7 b 36.1 ± 0.3 cd 31.3 ± 0.9 g

15

20 40 60

38.7 ± 2.5 cd 35.9 ± 1.0 cde 35.0 ± 3.4 cde

43.8 ± 1.3 d 43.5 ± 0.1 d 43.0 ± 0.9 e

37.3 ± 0.6 b 36.6 ± 0.1 c 35.5 ± 1.7 d

***

***

***

F-Test Temperature × duration

The data are presented as the means ± standard error, and 20 bulbs of each block were taken with three replications. Different letters in the same column indicate significant differences at the 0.05 level (ANOVA and LSD’s multiple range test), n = 3. CK stands for cloves stored continuously at ambient temperature (22 ± 2 ◦ C). *** Significant at 0.001 level (ANOVA and LSD’s multiple range test).

3.6. Interaction effects of low temperature and its duration treatment on garlic bulb character It was shown in Table 7 that the interaction of T × D significantly affected the bulb character reaching 0.001 level. The mean weight, diameter and height of the bulbs significantly decreased as a function of pre-planting clove chilling treatment (Table 7). The pattern was similar to the garlic bulb yield, as it decreased with the increase in the chilling duration. The control had the largest bulbs, followed by T5 D20 , while D60 had the smallest bulbs. These results indicate that longer chilling durations (D60 ) significantly reduced the mean weight, diameter and height of the bulbs. 4. Discussion Vernalization is critical to overwintering crops, as it prevents transition to the reproductive phase in regions with cold winters. As a plant that undergoes green plant vernalization, the vernalization of garlic requires a certain age of the plant, a low temperature and a specific duration. With the fulfillment of vernalization and

the subsequent appropriate climate conditions, the garlic will bolt. Therefore, the most important floral induction factors in garlic should be variety, bulb storage duration and temperature, and photoperiod and growth temperature (Pooler and Simon, 1993). Chilling treatment during bulb storage has been researched significantly on Allium and Lilium (Le Nard and De Hertogh, 1993; Bandara et al., 2000; Khokhar et al., 2007; Lee et al., 2007, 2008; Mojtahedi et al., 2012, 2013; Lucidos et al., 2014), but chilling treatment on bulbs as a substitute for plant vernalization has rarely been reported. The results of Karaguzel and Baktir (2013) indicate that cold treatments and their durations had significant effects on the flower stem length, flower stem diameter and flowering time of Allium robertianum Kolmann; the best results were obtained from bulbs stored at 10 ◦ C for a 30-day period in a heated glasshouse, while the earliest flowering was observed in bulbs stored at 5 ◦ C for 60 days in the glasshouse. Pooler and Simonn (1993) found that the flowering percentage of garlic was influenced most by clone, although interactions with photoperiod, growth temperature, and storage also occurred. Hypothetically, the clove, taken as a micro garlic plant, could sense the low temperature and saturate the vernalization response. Our results verified this hypothesis by showing

50

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

that the chilling treatment for a clove significantly stimulated the bolting of garlic and increased the bolting rate; the control plants that did not experience the low temperature only had a few bolts (Tables 5 and 6). This result implies that low-temperature treatment on a clove could substitute for the vernalization of the garlic plant. Cloves pre-treated under 10 ◦ C for 40-day or 5 ◦ C for 20day were recorded to have significantly higher bolting rates with a shorter growth period, which are very important for off-season cultivation. However, the lowest test temperature combined with the longest duration did not necessarily result in the highest bolting rate. This finding might be due to the weakened vegetative performance (highest secondary growth rate, leaves with dry yellow top and premature senescence) of the garlic plant under T5 D60 in the late growth stage (from 180 days after planting till harvest). The results further illustrated that much lower temperature would have resulted in better bolting performance, but too much lower temperature impeded the vegetative growth of garlic plant, which ultimately inhibited the subsequent bolting of garlic. Normally, the bolting rate of cv. G064 can reach 80–85% when fall-planted in the open field. Vernalization is saturated after going through low temperatures in the winter and early spring. However, this experiment was carried out to study the vernalization effect on the clove. Therefore, after the chilling treatment on the cloves (except for the control), the garlic plants were grown under 22 ± 2 ◦ C/16 ± 2 ◦ C (day/night) conditions throughout the experiment to eliminate the effect of low temperature during garlic growth (winter) on the results. In this manner, only the treated cloves received low temperature treatment, while the control was not subjected to low temperature (being stored continuously at ambient temperature (22 ± 2 ◦ C) and growing at 22 ± 2 ◦ C/16 ± 2 ◦ C (day/night) throughout). Without the fulfillment of vernalization, garlic plants only exhibited a 3.3% bolting rate (Tck ), whereas after the chilling treatment on the cloves, the bolting rate could reach 66.7% (T10 D40 ), which is in accordance with previous research and indicates that low temperature during garlic plant growth is indispensable for garlic bolting. It was proposed that a minimum critical cold period and low-temperature accumulation was needed for floral induction in garlic (Nakamura, 1985; Takagi, 1990; Pooler and Simon, 1993), with the optimum inducing condition differing among cultivars. Based on our results, the critical clove vernalization conditions for the bolting of cv. G064 are 10 ◦ C for 40 days. These results demonstrate that the chilling treatment on the cloves is effective and can satisfy the need for vernalization for bolting, which lays a strong basis for the developmental regulation of garlic production. Producers or farmers can produce fresh scapes or bulbs even in hot seasons or areas by treating the cloves with low temperature, which might enable the realization of a year-round supply of garlic to some degree. It was observed that plants under T5 or D60 had significantly greater plant standing height, especially T5 D60 and T10 D60 ; besides a significant influence of test temperature or its duration on plant standing height and leaf sheath length was obtained (except for main effect of temperature on plant standing height sampled 160 days after planting), while the significant effects was only recorded in early sampling (136 and/or 103 days after planting) when came to leaf sheath diameter, which indicated that chilling treatment promoted plant growth mainly through the enhanced plant elongation growth (Table 1). Seno et al. (1993) reported an increase in garlic plant height as a result of the increase in the vernalization duration from 40 to 60 days. These results are in agreement with those obtained in the present research. In contrast, Leal (1998) did not observe an effect of the vernalization period (25, 35 and 45 days) on the garlic plant standing height when evaluated at 53 and 90 days after planting. It was presumed that this fact was due to the climatic conditions of the pre-planting chilling treatment and the cropping location. Lucidos et al. (2014) showed that

low-temperature exposure affected stem elongation and leaf production in Lilium hansonii; exposure of bulbs to 4 ◦ C for 65 days promoted stem elongation and development of higher numbers of leaves with a 2-whorl leaf pattern. Kurtar and Ayan (2005) reported that exposure of tulips to low temperature increased the production of gibberellins and auxins, which are necessary for stalk elongation. According to Silva (1982), the greater garlic plant height was owing to the early plant emergence. At the start of growth, the plant relies on the clove for the canopy and root system development, which will later be an important source of reserve nutrient exportation, translating into fast growth. The low-temperature treatment on bulbs promoted the garlic plant standing height, which coincides with the results of other studies that showed that a low temperature might affect the plant growth promoters (carbohydrate content or plant growth regulators) that trigger early plant emergence and later fast growth (Lucidos, 2014). However, the advantage in the plant standing height of T5 D60 was only exhibited in the early stage of growth (from emergence till 170 days after planting) in this research. At the late growth stage (from 180 days after planting till harvest), the plants of T5 D60 turned yellow-green with drying on the top due to the high rate of secondary growth (data not shown). The secondary growth was significantly enhanced by lower-temperature and longer-duration treatments and was assumed to be one of the factors that promoted the plant’s premature senescence. The secondary growth occurred in T5 D60 , T5 D40 and T10 D60 , with the most severe and earliest occurrence observed in T5 D60 (data not shown). On the other hand, plants of T15 or D20 grew well throughout the whole growth period without any secondary growth. These results were in line with those reported by Ferreira et al. (1993) and Silva (1982), who found that the increase in the vernalization period resulted in a linear increase in the percentage of secondary growth of the garlic. Taken together, it is concluded that a low-temperature and long-duration treatment improved the plants’ elongation growth by triggering plant growth promoters, but too much low temperature combined with a longer duration (e.g., T5 D60 ) excessively stimulated plant growth and bud differentiation, resulting in an aggravated occurrence of secondary growth, which might lead to the weakened vegetative growth of the garlic plant in the late growth stage (from 180 days after planting till harvest). This can also explain the lower bolting rate of T5 D60 and T5 D40 . Our research illustrates that chilling treatment (including CK) reduced the plant growth period, mean bulb weight and bulb yield compared with plants grown in the open field under natural conditions (see Additional file 2 and Tables 5–7). The lower temperature or longer-duration chilling treatment had less yield and smaller garlic bulb compared with control. Cloves pre-treated under 5 ◦ C for 20 days or 10 ◦ C for 40 days showed superiority in bolting and bulbing ability with little difference from plants grown in the open field naturally compared with other chilling treatments. Though the effect of sole chilling treatment on cloves is not equal to that of green plant vernalization, the results provide evidence for the actual possibility of the vernalization application to produce garlic bulbs or fresh garlic scapes in warm season or area. Several researchers have reported the early maturity of garlic and onions after a pre-planting low-temperature treatment (Satti and Lopez, 1994; Khokhar et al., 2007). Bandara et al. (2000) found that increasing the chilling treatment period up to 45 days increased the bulb yield, while a further increase in the treatment period reduced the yield. However, there are many contradictions on the effect of the pre-planting low-temperature treatment on garlic yield; some researchers have reported increases (Ferreira et al., 1980), while others have observed no significant differences (Pyo et al., 1979); still others have reported depressive effects (Biasi and Mueller, 1984). The probable reason for the decrease in yield could be early ripening in longer cold treatments, which shortens the growth

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

period compared with shorter cold treatments. In the untreated control, where the cloves were kept at ambient temperature (20 ◦ C), the bulbs matured late. The findings agree with the research of Aura (1968), Butt (1968), Palilov (1969) and Khokhar et al. (2007) on onions; under low storage temperatures (0–7 ◦ C), the vegetative cycle is reduced and bulbing is accelerated, while under high temperatures (18–30 ◦ C), both bulbing and ripening are delayed. It is recognized that low temperature enhances the bulbing and cloving of garlic (Aoba and Takagi, 1971). Yamazaki et al. (2003) reported that plants exposed to low temperature (5 ◦ C for 35 days) did not induce the bulb formation of Allium X wakegi Araki but shortened the critical photoperiod for bulb formation, while bulbs stored at or below 15 ◦ C promoted the formation of new bulbs. Normally, small cloves planted under adverse environmental conditions would produce undesirable one-clove bulbs, while suitable planting environments will give rise to normal bulbs. In the present study T5 D40 and T5 D60 were observed with weakened growth due to premature senescence, which accounted for the increase in the undesirable one-clove bulbs. Furthermore, the control (with a longer growth period) grown under warm conditions had the highest yield and largest number of normal bulbs (no one-clove bulbs). The results raised doubt about the necessity of pre-plant chilling for bulbing. Experiments are needed to further demonstrate this issue. It is reported that flower induction and development in garlic is a complex process and depends not only on environmental factors but also on the genetic background (Pooler and Simon, 1993). As garlic is a very variable crop, the obtained results (critical conditions for clove vernalization treatment) are relevant for a specific cultivar (cv. G064) in a specific region (subtropical monsoon climate). Further work should explore the vernalization requirement of non-bolting or semi-bolting cultivars and determine if more cultivars could be induced to bolt under our research conditions. Nevertheless, the findings provide strong evidence for the cultivation of garlic of various cultivars in different climates or seasons and makes off-season cultivation possible. Researchers could use a similar methodology (treating garlic cloves with cold temperature for certain durations before planting) to shorten the days needed for harvest or even to produce desirable bolts and bulbs in warm seasons.

5. Conclusion The results of this study show that the growth and development of the garlic plant significantly changed in response to the clove chilling treatment. The lower temperature significantly improved the elongation growth and earliness of the plant, promoted the oneclove bulb rate, stimulated the bolting and increased the bolting rate but decreased the garlic bulb yield and mean bulb weight. This study verified that low-temperature treatment on the clove could act as a vernalization treatment for garlic plants and contradicted the classic view of chilling (lower than 20 ◦ C) as being essential for cloving and bulbing of garlic based on the results that garlic plants even without exposure to low temperature (lower than 20 ◦ C) gave rise to normal bulbs. Further researches are needed to deeply elaborate this question. Generally speaking, cloves treated at 5 ◦ C for 20 days or 10 ◦ C for 40 days before planting tend to be suitable for off-season cultivation of cv. G064 with a relatively short growth period and a high bolting rate and yield. Once the vernalization requirement of a specific cultivar is known, it would be possible for a producer or breeder to plan a sound production program.

51

Acknowledgements This research was supported by a State Commonwealth (Agriculture) Scientific Research project (No. 200903018) and a State Natural Science Foundation project (No. 31171949).

Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.scienta.2015. 07.018

References Aoba, T., Takagi, H., 1971. Studies on the bulb formation in garlic plants III. On the effects of cooling treatments of seed-bulbs and day-length during the growing period on bulbing. J. Jpn. Soc. Hortic. Sci. 40 (3), 40–45. Aura, K., 1968. Studies on the vegetatively propagated onions cultivated in Finnland, with special reference to flowering and storage. IX. The influence of various storage temperatures on flowering and yield in the North Finnish onion strain. Ann. Agric. Fenn. 7, 183–188. Bandara, M.S., Krieger, K., Slinkard, A.E., Tanino, K.K., 2000. Pre-plant chilling requirements for cloving of spring-planted garlic. Can. J. Plant Sci. 80 (2), 379–384. Biasi, J., Mueller, S., 1984. Influência da imersão do alho em água. In: Jaboticabal, S.P. (Ed.), C Ongresso Brasileiro De Olericultura, 24. Resumos, Brasília, p. 69. Butt, A.M., 1968. Vegetative Growth, Morphogenesis and Carbohydrate Content of the Onion Plant as a Function of Light and Temperature under Field and Controlled Conditions, 68. Mededelingen Landbouwhogeschool Wageningen, pp. 94–191. Cheng, Z.H., Lu, G.Y., Du, S.L., 1991. Studies on the concept and classification of secondary growth in garlic plants. Acta Hortic. Sinica 18 (4), 345–349. Dong, Y.X., Cheng, Z.H., Meng, H.W., Liu, H.Q., Wu, C.N., Khan, A.R., 2013. The effect of cultivar, sowing date and transplant location in field on bolting of welsh onion (Allium fistulosum L.). BMC Plant Biol. 13, 154. Etoh, T., Simon, P.W., 2002. Diversity, fertility and seed production of garlic. In: Rabinowitch, H.D., Currah, L. (Eds.), Allium Crop Sciences: Recent Advances. CAB International, Wallingford, pp. 101–117. Ferreira, F.A., Casali, V.W.D., Álvares V.H., Rezende, G.M., 1993. Desenvolvimento de alho, cultivares Chonan e Quitéria, após armazenamento refrigerado, in: EMPRESA DE PESQUISA AGROPECUÁRIA DE MINAS GERAIS, Projeto olericultura: relatório de pesquisa 1987/92. Belo Horizonte, Epamig, pp. 28–30. Filgueira, F.A.R., 2008. New Manual of Olericulture, Third ed. UFV, Vic¸osa, pp. 421. Kamenetsky, R., Rabinowitch, H.D., 2001. Floral development in bolting garlic. Sex Plant Reprod. 13, 235–241. Kamenetsky, R., Shafir, I.L., Baizerman, M., Khassanov, F., Kik, C., Rabinowitch, H.D., 2004a. Garlic (Allium sativum L.) and its wild relatives from Central Asia: evaluation for fertility potential. Acta Hortic. 637, 83–91. Kamenetsky, R., Shafir, I.L., Zemah, H., Barzilay, A., Rabinowitch, H.D., 2004b. Environmental control of garlic growth and florogenesis. J. Am. Soc. Hortic. Sci. 129, 144–151. Karaguzel, O., Baktir, I., 2013. The effects of cold treatments on the growth and flowering characteristics of Allium robertianum Kolmann bulbs. J. Food Agric. Environ. 11 (2), 1174–1177. Khokhar, K.M., Hadley, P., Pearson, S., 2007. Effect of cold temperature durations of onion sets in store on the incidence of bolting, bulbing and seed yield. Sci. Hortic. 112, 16–22. Kurtar, E.S., Ayan, A.K., 2005. Effects of gibberellic acid (GA3) and indole acetic acid (IAA) on flowering, stalk elongation and bulb characteristics of tulip (Tulipa gesneriana var. Cassini). Pak. J. Biol. Sci. 8, 273–277. Le Nard, M., De Hertogh, A.A., 1993. Bulb growth and development and flowering. In: De Hertogh, A.A., Le Nard, M (Eds.), The Physiology of Flowering Bulbs. Elsevier, Amsterdam. Leal, F.R., 1998. Períodos de hidratac¸ão, vernalizac¸ão, cobertura morta e material orgânica, sobre as características agronômicas da cultura do alho, cultivar Roxo Pérola de Cac¸ador. Jaboticabal, UNESP-FCAV, pp.132 (Tese doutorado). Lee, A.K., Suh, J.K., Roh, M.S., 2007. Optimum bulb vernalization temperature of Asiatic hybrid lilies, Lilium x elegans Thunb. Hortic. Environ. Biotechnol. 48, 303–313. Lee, A.K., Suh, J.K., Roh, M.S., 2008. Requirement of bulb vernalization and shoot photoperiod for flowering of Lilium x elegans cultivars. Hortic. Environ. Biotechnol. 49, 90–97. Lucidos, J.G., Younis, A., Hwang, Y.J., Son, B.G., Lim, K.B., 2014. Determination of optimum conditions for breaking bulb dormancy in relation to growth and flowering in Lilium hansonii. Hortic. Environ. Biotechnol. 55 (4), 257–262. Mojtahedi, N., Masuda, J., Hiramatsu, M., Okubo, H., 2012. Latitudinal variation of bulb dormancy and early flowering ability in Lilium longiflorum and Lilium formosanum populations of the Ryukyu Archipelago and Taiwan. In: Proc. of XI Inter. Sym. on Flower Bulbs and Herbaceous Perennials, 28 March–01 April, 2012, Antalya, Turkey, p. 48.

52

C. Wu et al. / Scientia Horticulturae 194 (2015) 43–52

Mojtahedi, N., Masuda, J.I., Hiramatsu, M., Hai, N.T.L., Okubo, H., 2013. Role of temperature in dormancy induction and release in one-year-old seedlings of Lilium longiflorum populations. J. Jpn. Soc. Hortic. Sci. 82, 63–68. Motaz, B.M., Omar, F.A., Abd Elkader, E.L., Shiaty, M., Imam, A.A., Galm, G., Shahin, H.A., Zein, A., 1971. The effect of some treatment on yield and quality of Egyptian garlic. Agr. Rev. 49, 157–172. Nakamura, E., 1985. Allium. In: Halevy, A.H. (Ed.), CRC Handbook of Flowering, vol. I. CRC Press, Boca Raton, FL, pp. 410–418. Palilov, N.A., 1969. The biological bases of onion storage. Proceedings of the Symposium of vegetable storage, Acta Horticulturae 20, 53–64. Pooler, M.R., Simon, P.W., 1993. Garlic flowering in response to clone, photoperiod, growth temperature, and cold storage. HortScience 28 (11), 1085–1086. Pyo, H.K., Lee, B.Y., Moon, W., Woo, J.K., 1979. A study of the development of a new cultural system for garlic (1). The effect of lowtemperature bulb treatment, night interruption and supplementary lighting on the growth and bulbing of garlic in a plastic film house. J. Korean Soc. Hortic. Sci. 20, 19–27. Resende, J.T.V., Morales, R.G.F., Resende, F.V., Faria, M.V., Souza, R.J., Marchese, A., 2011. Garlic vernalization and planting dates in Guarapuava. Hortic. Bras. 29, 193–198. Ritz, C., Pipper, C., Yndgaard, F., Fredlund, K., Steinrucken, G., 2010. Modeling flowering of plants using time-to-event methods. Eur. J. Agron. 32 (2), 155–161. Satti, S.M.E., Lopez, M., 1994. Effects of storage-temperature on growth and bulb formation in 4 garlic (Allium sativum L.) cultivars. Pak. J. Bot. 26 (1), 161–165.

Sendl, A., 1995. Allium sativum and Allium ursinum: part 1. Chemistry analysis, history, botany. Phytomedicine 4, 323–339. Seno, S., Castellan, P.D., Kimoto, T., 1993. Influência do tempo de vernalizac¸ão e da época de plantio na cultura do alho (Allium sativum L), Cv. Roxo Pérola de Cac¸ador, na região de Ilha Solteira-SP. Científica 21, 275–285. Siddique, M.A., Rabbani, M.G., 1985. Growth and bulbing of garlic in response to low temperature treatment of bulb and planting date. Bangladesh J. Bot. 14, 41–46. Silva, J.L.O., 1982. Análise de crescimento de alho (Allium sativum L.) cultivar Chonan, sob três períodos de frigorificac¸ão pré-plantio dos bulbos. Lavras, ESAL, pp.76 (Tese mestrado). Song, S.R., Han, S., Zhang, E.R., 2010. Alliaceous vegetables. In: Cheng, Z.H. (Ed.), Systematics of Olericulture Science. Science Press, China, pp. 187–195. Takagi, H., 1990. Garlic Allium sativum L. In: Rabinowitch, H.D., Brewster, J.L. (Eds.), Onions and Allied Crops, Vol. 3 (biochemistry, Food Science, and Minor Crops). CRC Press, Boca Raton FL, pp. 109–157. Trevaskis, B., Hemming, M.N., Dennis, E.S., Peacock, W.J., 2007. The molecular basis of vernalization-induced flowering in cereals. Trends Plant Sc. 12 (8), 352–357. Yamazaki, H., Hamano, M., Yamato, Y., Miura, H., 2003. Bulbing response of Allium X wakegi Araki to temperature experienced prior to bulb formation. J. Jpn. Soc. Hortic. Sci. 72 (1), 69–74.