Abstract
Pseudobulbs are carbohydrate storage organs in Oncidesa. A current pseudobulb forms on a developing vegetative shoot in each growth cycle and it becomes a back pseudobulb when the next vegetative shoot emerges. Both current and back pseudobulbs store carbohydrates, but their functions might differ because the inflorescence emerges from the new (current) shoot after the shoot has developed to a certain stage. This study investigated carbohydrate storage and use in current and back pseudobulbs. We analyzed carbohydrates in the current pseudobulb at five stages during inflorescence development. Glucose and fructose were the highest in the current pseudobulb in the first two stages, when the inflorescence was 10 to 35 cm tall. Then, both glucose and fructose decreased in the following stages to support inflorescence development, but starch increased at that time. In addition, we used Oncidesa with one or two new vegetative shoots to study the use of carbohydrates in pseudobulbs during growth cycles. In both plants with one or two shoots, glucose and fructose accumulated when current pseudobulbs formed, but plants with two new shoots had smaller current pseudobulbs and lower monosaccharide concentrations. Plants with two shoots also consumed more starch in all back pseudobulbs, whereas in the plants with one new shoot, starch only decreased significantly in the first back pseudobulb, which was closer to the new shoot. In addition, if an inflorescence did not develop in the previous growth cycle, new shoots used the monosaccharides that remained in the youngest back pseudobulb for growth; at the same time, starch accumulated in all back pseudobulbs. The current pseudobulb was the actively growing part. Its main carbohydrates were monosaccharides, which accounted for 25% of dry weight and Oncidesa used these carbohydrates mainly for inflorescence growth. After monosaccharides in the pseudobulb were used, the pseudobulb began to store starch. Back pseudobulbs, in which >50% of dry weight was starch, were the primary storage organs that supported new vegetative shoot growth and partly supported later inflorescence development that emerged from the new (current) shoot.
Oncidesa is a man-made genus by crossing Oncidium and Gomesa, which is an important cut-flower crop and a sympodial orchid with an enlarged pseudobulb on each shoot. In the developing vegetative shoot (current shoot), an inflorescence bud forms before pseudobulb expands and then inflorescence elongates when the pseudobulb is nearly fully expanded.
However, the industry is facing a problem in that some plants do not bloom and the flowering mechanism of Oncidesa has not been well studied. Higher light intensity increased the quality of the inflorescence and a shaded environment inhibited inflorescence elongation of Oncidesa Gower Ramsey (Chang and Lee 1998). A low temperature (13 to 20 °C) increased inflorescence quality but delayed flowering time compared with a high temperature (20 to 30 °C) in Oncidesa Gower Ramsey (Chang and Lee, 2000). However, temperatures >35 °C inhibited the spiking (Chin et al. 2014). Environmental factors would accelerate flowering, but not the necessary flowering signals for Oncidesa. Therefore, there might be other factors that inhibit flowering. The nutrient status might determine the flowering ability of Oncidesa because it flowers at the end of each growth cycle regardless of the seasons. Nevertheless, it is difficult to investigate the relationship between flowering and carbohydrate storage in Oncidesa, which has multiple pseudobulbs as storage organs.
Carbohydrate availability also affected the flowering quality of Oncidesa. The current pseudobulb and back pseudobulbs were all important carbohydrate sources for flowering of Oncidesa Goldiana (Yong and Hew 1995a). The current pseudobulb of Oncidesa Gower Ramsey accumulated a large amount of glucose, fructose, and mannan and those sugars decreased during the inflorescence development (Wang et al. 2003, 2008). The photosynthetic rate of the leaf on the node bearing inflorescence increased during the inflorescence development in Oncidesa Goldiana, which is a strategy to increase carbohydrate availability during flowering (Hew and Yong 1994).
There is no detailed study on the carbohydrate form or metabolism in back pseudobulbs, but when there were more back shoots connected to the current shoot, the quality of inflorescence improved in Oncidesa Goldiana (Yong and Hew 1995a). The length, branch number, and floret count of inflorescence increased when the number of back shoots increased from zero to two; however, when the number of back pseudobulbs was more than two, the quality of inflorescence did not increase further (Yong and Hew 1995a). Isotope 14C tracing also showed that >60% of the photosynthate produced by back shoots was transported to the inflorescence on the current shoot in Oncidesa Goldiana (Yong and Hew 1995a). In other sympodial orchid species, such as Catasetum viridiflavum and Dimerandra emarginata, growth and flowering quality were also improved when the number of back pseudobulbs increased, which showed the influence of the carbohydrates from back pseudobulbs (Zimmerman 1990; Zotz 1999). In a word, both current and back pseudobulbs contributed carbohydrates to the reproductive growth and increased flowering quality.
The flowering process of plants is divided into flower initiation and flower development. Flower initiation is often related to environmental factors or autonomous factors like plant age or hormones (Corbesier and Coupland 2006). Some studies indicated that carbohydrate availability was also a limitation of flower initiation (Ponnu et al. 2011; Wahl et al. 2013). Flower development required sufficient carbohydrate supplements (Bernier 1988). However, it is not understood fully that the inhibition of the flowering process in Oncidesa occurs during flower initiation or development.
Oncidesa has a juvenile stage of ∼1.5 to 2 years and, after the juvenile stage, it flowers at the end of each growth cycle after the current pseudobulb expands entirely. However, the flower initiation time of Oncidesa indicates that there are different roles for current and back pseudobulbs in reproductive growth. Gomesa Boissiense formed nodes and branch primordia of the inflorescence before the current pseudobulb expanded, which indicated that the flowers were initiated before carbohydrates accumulated in the current pseudobulb (Tanaka et al. 1986). If flower initiation was related to carbohydrate availability, it would be controlled by the carbohydrates in back pseudobulbs and the carbohydrates accumulated later in the current pseudobulb might contribute to the growth and elongation of the inflorescence.
The information about the carbohydrates in back pseudobulbs of Oncidesa is limited. We conducted this study to understand carbohydrate changes in both the current and back pseudobulbs during growth cycles and to investigate the roles of different pseudobulbs in the development of Oncidesa plants.
Materials and Methods
Expt. I. Changes in carbohydrates in the current pseudobulb during inflorescence growth
Plant materials.
Current shoots of Oncidesa Gower Ramsey ‘Honey Angel’ with inflorescences at five stages were collected at an orchid nursery in Taichung, Taiwan, on 17 Sep 2018. Stages 1 to 5 were defined as inflorescence length at 10 cm, 35 cm, 65 cm, inflorescence well-branched with unopened flower buds, and 50% florets opening. Each stage had 10 single-plant replications.
Data collection.
Plants were collected in the nursery in the morning and were then shipped to the laboratory on the same day. We first rinsed the plants with water to remove the dirt and separated the current shoot into leaves, pseudobulb, and inflorescence. For the plants at stages 4 and 5, we further divided the inflorescence into stalks and florets (unopened buds and opened florets). The fresh weight of each part was measured, and the dry weight and carbohydrate concentration were determined after freeze drying.
Expt. II. Changes in carbohydrates in current and back pseudobulbs during vegetative growth
Plant materials.
Forty-two plants of Oncidesa Gower Ramsey ‘Honey Angel’ were collected in a nursery in Taichung, Taiwan, on 20 Oct 2018. Half of them had one new vegetative shoot 10 cm in height and the other half had two new shoots that were 10 cm in height. All plants were transplanted into 13.5-cm plastic pots in a medium of 1 gravel:1 charcoal. If there were more than two back shoots, the excess back shoots were trimmed. The size and number of the back pseudobulbs were equal among plants to ensure that all had similar conditions.
Plants were grown in a plastic greenhouse with a pad and fan cooling system and a shade net at National Taiwan University, Taipei, Taiwan. Plants were fertigated with a 15N–2.1P–12.4K water-soluble fertilizer (Peters Excel 15–5–15 Cal–Mg; ICL Specialty Fertilizers, Dublin, OH, USA) at 100 mg·L−1 N twice a month, and 0.5 g of controlled release fertilizer 13N–4.3P–9.1K (Hi-Control, S101, 13–11–10–2TE, Type 100; Asahi Kasei, Tokyo, Japan) was added to the medium every 100 d. Irrigation and pesticides were applied when needed. In the first growth cycle, 0 to 150 d after transplanting (DAT), the average daytime photosynthetic photon flux density (PPFD) was 190 μmol·m−2·s−1 [daily light integral (DLI) was 7.9 mol·m−2·d−1] and the average temperature was 20 °C. In the second growth cycle, 151 to 300 DAT, the average daytime PPFD was 230 μmol·m−2·s−1 (DLI was 10.2 mol·m−2·d−1) and the average temperature was 27 °C.
The terms “current shoot” and “first back shoot” represent the developing new shoot and the shoot that has developed in the last growth cycle in sympodial orchid. The ordinal number of each shoot changed as the plant growth, for example, the current shoot at the beginning of this experiment became the first back shoot at the end. Therefore, we labeled the shoots from the oldest to youngest as A to D shoot. At the beginning of the experiment, the second and first back shoots and the new shoot(s) were labeled as A shoot, B shoot, and C shoot(s), respectively (Fig. 1). The plants with two new vegetative shoots possessed two C shoots. The first growth cycle was from C shoot at 10 cm height until the next vegetative shoot emerged, and this newly emerged shoot was then labeled as a D shoot. The C shoot did not flower in the experiment. The second growth cycle was from the emergence of the D shoot until an inflorescence emerged on the D shoot. At the end of the second growth cycle, A, B, and C shoots were considered the third, second, and first back shoots, respectively (Fig. 1).
Schematic of Oncidesa Gower Ramsey ‘Honey Angel’ with one or two new shoots. Plants were sampled at the end of 0th, first, and second growth cycle. The shoot was labeled as shoot A to shoot D depending on the order of shoot age; shoot A was the oldest shoot.
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
Biomass measurements.
Shoot length and pseudobulb volume of C and D shoots were measured every week. Plants were sampled at three stages (Fig. 1), which were the end of the 0th growth cycle (10 DAT, C shoots at 10 cm), the end of the first growth cycle (150 DAT, D shoot at 10 cm) and the end of the second growth cycle (300 DAT, the inflorescence on the D shoot reached 10 cm). Because all plants did not flower at the end of the first growth cycle, we collected plants when the D shoots had just emerged. We collected seven Oncidesa with one or two shoots each and we dissected the plants into various parts. Fresh and dry weights were measured and then the carbohydrate concentrations of each pseudobulb were analyzed. There were two C shoots and two D shoots on the plants with two shoots; therefore, shoot length, pseudobulb volume, dry weight, fresh weight, and carbohydrates of C and D shoots on the plants with two shoots were shown as the average of two shoots.
Carbohydrate analysis.
In both experiments, all samples were collected in the morning and fixed in liquid nitrogen for a consistent result. Samples were freeze-dried and ground into powder. For soluble sugars, we weighed 50 mg of sample fine powder into tubes to which 100 μL of 1% raffinose was added as an internal standard. The sample was extracted at 70 °C with 80% ethanol, three extractions of 3 mL each, 30 min per extraction. The supernatants of these extractions were collected and combined, purified using an ion-exchange resin column with 1 mL anion exchange resin (Amberlite IRA-67, acetate form; Sigma-Aldrich, St. Louis, MO, USA) and 1 mL cation exchange resin (Dowex-50W, hydrogen form; Sigma-Aldrich). Finally, we used a vacuum evaporator (Labconco Co., Kansas City, MO, USA) to reduce the sample to less than 1 mL and then we diluted it to 10 mL with Milli-Q water.
The leftover residue contained certain water-soluble polysaccharides (WSPs) and water-insoluble starch. We extracted WSPs following the method of Ranwala and Miller (2008). The residue was extracted using Milli-Q water, in which CaCO3 was used to adjust the pH to 8. The extraction was conducted twice at 70 °C in a water bath for 30 min each. Then, the supernatant was hydrolyzed with 1 N HCl for 3 h.
For starch analysis, we used 30 mg potato starch (Merck, Darmstadt, Germany) as an external standard. After extracting soluble sugar and WSPs, we gelatinized the residue in a 100 °C water bath for 6 h with 2 mL Milli-Q water in the tube. After cooling, 2 mL of starch-degrading enzymes α-amylase (100 units per sample; Sigma-Aldrich), pullulanase (9 units per sample; Sigma-Aldrich), and amyloglucosidase (10 units per sample; Sigma-Aldrich) in pH 4.7 sodium citrate buffer solution were added to break down the starch into simple sugars at 37 °C for 6 h, followed by treatment at 55 °C for 18 h. Then, we centrifuged the sample and collected the supernatant. The supernatant was diluted to 10 mL with Milli-Q water for analysis.
To quantify carbohydrates, the extractants were subjected to high-performance anion exchange chromatography with pulsed amperometric detection (HPAE-PAD; Dionex Co, Sunnyvale, CA, USA). The chromatography system consisted of a GS50 gradient pump, an ED50 detector, and a CarboPac PA-1 analytical (4
Statistical analysis.
Two experiments used a completely randomized design. Mean separation for multiple comparison was carried out by the least significance difference test at P ≤ 0.05 when the P value of analysis of variance ≤ 0.05. A Student’s t test at P ≤ 0.05 was used for mean separation of paired comparison. We used Costat (Version 6.1; CoHort Software, Monterey, CA, USA) for statistical analysis and SigmaPlot software (Version 10.0; Systat Software, San Jose, CA, USA) to plot figures.
Results
Expt. I. Changes in carbohydrates in the current pseudobulb during inflorescence growth.
The dry weight of the current pseudobulb continued to increase after inflorescence had emerged. When the inflorescence developed from 10 to 35 cm, the dry weight of current shoot increased from 2.19 to 2.49 g (Table 1). The fresh weight of pseudobulb maintained at 34.7 to 41.4 g and dropped to 31.7 g when 50% of the florets opened (Table 1). Both fresh and dry weights of the inflorescence increased as it developed (Table 1). The water content of the pseudobulb was >90% and decreased after florets opened, whereas the water content of the florets increased significantly at the same time (data not shown).
Fresh weight and dry weight of various parts of the current shoot of Oncidesa Gower Ramsey ‘Honey Angel’ during different stages of inflorescence development.
During the first two stages of inflorescence development, the current pseudobulb had the highest soluble sugar concentrations of all stages (Fig. 2A). Glucose concentration was 130 mg·g−1 and fructose concentration was 129 mg·g−1 at stage 1 (Fig. 2A). As the inflorescence elongated, the glucose and fructose concentration decreased. At stage 5, the glucose and fructose concentration decreased to 60 and 53 mg·g−1, respectively (Fig. 2A). In comparison, sucrose concentration in the current pseudobulb maintained a low and stable level at 15 to 19 mg·g−1 throughout inflorescence development (Fig. 2A).
Changes in soluble sugars (A) and reserve carbohydrates (B) concentrations in the current pseudobulb of Oncidesa Gower Ramsey ‘Honey Angel’ during inflorescence development. Stages 1 to 5 means inflorescence length at 10 cm, 35 cm, 65 cm, inflorescence well-branched with unopened florets, and 50% florets opening, respectively. Bars indicate standard error of the mean. Different letters separate the means among the stages of each carbohydrate by least significant difference test (P ≤ 0.05; n = 10) and ns represents the means are not significantly different among the stages by analysis of variance (n = 10).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
The concentrations of reserve polysaccharides were low relative to soluble sugars in the current pseudobulb. The change in WSPs concentration was similar to soluble sugars, but it was only 27 mg·g−1 at stage 1 (Fig. 2B). However, the shift in starch concentration was the opposite of WSPs and soluble sugars. The concentration of starch increased from 30 to 60 mg·g−1 during the development of inflorescence, but this was not significant statistically (Fig. 2B). The main WSP in the current pseudobulb of Oncidesa was mannan. The composition of WSPs included arabinose, galactose, glucose, mannose, and fructose, in which mannose accounted for >80% of sugar units of WSPs (Fig. 3).
Changes in the content of sugar units in water-soluble polysaccharides (WSPs) in the current pseudobulb of Oncidesa Gower Ramsey ‘Honey Angel’ during inflorescence development. The sugar units were hydrolyzed from WSPs. Stages 1 to 5 means inflorescence length at 10 cm, 35 cm, 65 cm, inflorescence well-branched with unopened florets, and 50% florets opening, respectively. Bars indicate standard error of the mean. Different letters separate the means among the stages of each carbohydrate by least significant difference test (P ≤ 0.05; n = 10) and ns represents the means are not significantly different among the stages by analysis of variance (n = 10).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
Expt. II. Changes in carbohydrates in current and back pseudobulbs during vegetative growth.
The C shoot (new shoot) elongated rapidly in the early stage of the first growth cycle and the elongation rate decreased after 50 DAT (Fig. 4A). The plants entered the unsheathing stage at ∼90 DAT, and both the size of pseudobulb and the shoot length reached a maximum on 150 DAT (Fig. 4A). At the same time, D shoot had emerged for 30 d and reached 5 cm in length (Fig. 4B). After 150 DAT, the elongation rate of the D shoot increased (Fig. 4B) and the volume of pseudobulb on shoot C (referred to as C pseudobulb hereafter) decreased (Fig. 4A), the plants entered the next growth cycle. The length of C shoot showed no difference between plants with one or two vegetative shoots, although the pseudobulb volume of plants with two shoots was smaller (P = 0.02; Fig. 4A). The pseudobulbs in plants with two vegetative shoots also shrank faster than those in plants with only one vegetative shoot after it became a back pseudobulb (Fig. 4A). The D pseudobulb appeared on ∼230 DAT and reached a maximum volume on 300 DAT (Fig. 4B). The maximum volume of D pseudobulb in plants with two shoots was also smaller than that of the plants with one shoot (P = 0.008; Fig. 4B).
Effects of new shoot numbers on the shoot length and pseudobulb volume of C shoot (A) and D shoot (B) of Oncidesa Gower Ramsey ‘Honey Angel’. C shoot was the new developing shoot at the beginning of the experiment and D shoot was the vegetative shoot that emerged on the mature C shoot. Data for plants with two shoots (two C shoots) are averages (n = 14 during C shoot growth and n = 7 during D shoot growth).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
The fresh weight of back pseudobulbs decreased with growth cycles. The fresh weight of A pseudobulb of plants with one and two shoots fell from 23.7 to 23.3 g and from 37.3 to 19.7 g, respectively (Table 2). B pseudobulb and C pseudobulb also exhibited the same trend with growth cycles. The fresh weight of the youngest pseudobulb was the highest in a plant, such as B pseudobulb in the 0th growth cycle, C pseudobulb in the first growth cycle, and D pseudobulb in the second growth cycle (Table 2). Plants with one shoot had a higher D pseudobulb fresh weight of 45.5 g than plants with two shoots of 27.8 g (Table 2). The dry weight difference among growth cycles and shoot order was not as obvious as fresh weight (Table 3), which showed that there was a significant change in water content in pseudobulbs, where the plant stored water and contributed this water to new growth as pseudobulbs aged. In plants with two vegetative shoots, dry weights of C and D pseudobulbs were lower than those of A and B pseudobulbs in the second growth cycle (Table 3); however, the fresh weights of C and D pseudobulbs were higher than those of A and B pseudobulbs because their water storage had not been consumed yet (Table 2). There was also a difference in dry weight between plants with one or two shoots. During the second growth cycle, the average dry weight of C and D pseudobulbs was lower than that of plants with one shoot (Table 3).
Changes in fresh weight of each pseudobulb of Oncidesa Gower Ramsey ‘Honey Angel’ with one or two new shoots during two growth cycles.
Changes in dry weight of each pseudobulb of Oncidesa Gower Ramsey ‘Honey Angel’ with one or two new shoots during two growth cycles.
The soluble sugars in A and B pseudobulbs remained relatively low during growth and there was no difference between plants with one or two shoots (Fig. 5D and H). Concentrations of glucose and fructose were 10 to 30 mg·g−1 and sucrose concentration remained at 30 mg·g−1 in A and B pseudobulbs throughout the experiment (data not shown). From the 0th growth cycle to the first growth cycle, C pseudobulb developed into fully mature current pseudobulb. The swelling C pseudobulb accumulated a high level of monosaccharides, glucose, and fructose, but the level of sucrose was similar to that of A and B shoots (Fig. 5). Glucose and fructose contents in the C pseudobulb were ∼400 mg (Fig. 5A, B, E, and F) and the concentrations of glucose and fructose were ∼110 mg·g−1 (data not shown). During the second growth cycle, C shoot became the first back shoot, and glucose and fructose in C pseudobulb decreased (Fig. 5A, B, E, and F). Although C shoot did not flower during the experiment, the sugars in C pseudobulb were used in the vegetative growth of D shoot instead of the inflorescence. D pseudobulb also accumulated a high level of glucose and fructose during the second growth cycle when it turned into a mature current shoot, which was similar to the C pseudobulb during the first growth cycle (Fig. 5A, B, E, and F). However, glucose (P = 0.019), fructose (P = 0.012), sucrose (P = 0.045), and total soluble sugar contents (P = 0.013) in D pseudobulbs of plants with two vegetative shoots (Fig. 5E–H) were significantly lower than those of plants with one vegetative shoot (Fig. 5A–D).
Changes in soluble sugar contents in Oncidesa Gower Ramsey ‘Honey Angel’ with one new shoot (A–D) or two new shoots (E–H) during various growth cycles. Bars indicate standard error of the mean (n = 7).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
The changes in WSPs in pseudobulbs (Fig. 6) were similar to those of soluble sugars. WSPs were higher in the current pseudobulb, such as C pseudobulb in the first growth cycle and D pseudobulb in the second growth cycle (Fig. 6). The concentration of WSPs was lower than soluble sugars and was ∼60 mg·g−1 in C pseudobulb in the first growth cycle and 40 mg·g−1 in D pseudobulb in the second growth cycle. Both concentration and content of WSPs in D pseudobulb were not statistically different in plants with one shoot (Fig. 6A and B) compared with those with two shoots (Fig. 6C and D).
Changes in water-soluble polysaccharides (WSPs) of Oncidesa Gower Ramsey ‘Honey Angel’ with one new shoot (A–B) and two new shoots (C–D) during various growth cycles. Bars indicate standard error of the mean (n = 7).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
At the 0th growth cycle, the concentrations of starch in A and B pseudobulbs of plants with one shoot were 572 and 324 mg·g−1, respectively, which meant that >50% of dry matter in A pseudobulb was starch (Fig. 7A). During the first growth cycle, only the starch in B pseudobulb of plants with one shoot decreased (Fig. 7B), the starch decreased in both A and B pseudobulbs of plants with two shoots (Fig. 7D). Because there was no decrease in starch in A pseudobulb of plants with one shoot (Fig. 7B), the starch content of plants with one shoot was higher than plants with two shoots (P= 0.04; Fig. 7B and D). The C pseudobulb was the current pseudobulb during the first growth cycle with a low starch level (Fig. 7); after it became a back pseudobulb during the second growth cycle, the starch level increased. In addition, the increase in starch content of the plants with one shoot was higher than that of plants with two shoots during the second growth cycle (P = 0.02; Fig. 7B and D).
Changes in starch in Oncidesa Gower Ramsey ‘Honey Angel’ with one new shoot (A–B) and two new shoots (C–D) during various growth cycles. Bars indicate standard error of the mean (n = 7).
Citation: HortScience 58, 1; 10.21273/HORTSCI16905-22
Discussion
In this study, the current pseudobulb of Oncidesa continued to develop and accumulated dry matter at the early stages of inflorescence development (Table 1). By calculating the change in fresh weight and dry weight of the current pseudobulb during inflorescence growth, we found a significant decrease in water content in the current pseudobulb when the florets opened (Table 1). The decrease in fresh weight in back pseudobulbs (A and B pseudobulbs) was also found during growth cycles (Table 2), although dry weight barely changed (Table 3), demonstrating the water storage function of pseudobulbs and the use of water for vegetative and reproductive growth.
The current pseudobulb had the highest level of carbohydrate at the early stages of inflorescence development (Fig. 2A); at the same time, current pseudobulb reached its maximum weight (Table 1). The main carbohydrates in current pseudobulb were monosaccharides (glucose and fructose), which accounted for 26% (130 + 129 mg·g−1) of dry weight (calculated from Fig. 2A). The reserve polysaccharides, which included starch and WSPs, only accounted for 6% of dry weight in the current pseudobulb (Fig. 2B). Wang et al. (2003) found that the soluble sugar concentration was 30% in the current pseudobulb, which was similar to our results.
Soluble sugars decreased significantly in the current pseudobulb when the inflorescence grew (Fig. 2A), which demonstrated that soluble sugars in the current pseudobulb supported the growth of inflorescence. Yong and Hew (1995a) reported that 52% of photosynthate in the inflorescence came from the current shoot and the other 48% came from multiple back shoots. Although back shoots provided part of the carbohydrates, the inflorescence used more carbohydrates supplied by the current shoot.
In addition, pure mannan, which is a WSP, was detected in current pseudobulbs in Oncidesa (Wang et al. 2006). Mannan was one type of hemicellulose in the cell wall (Scheller and Ulvskov 2010) and was not the energy source in most scenarios. However, in some cases, the mannan in the seed cell wall degraded to supply energy for germination (Buckeridge 2010). Mannan was also found in many bulbous crops as an energy source, which accounted for 1% to 14% of dry weight (Ranwala and Miller 2008). In this study, we also found mannan in the current pseudobulb of Oncidesa, but in a relatively low concentration (Fig. 6).
It is widely accepted that flowering in Oncidesa is controlled by nutrient status. However, it seems that flower initiation is not related to the carbohydrates in the current pseudobulb in Oncidesa, even though it contributed a significant amount of soluble sugars to the inflorescence late in the growth cycle (Fig. 2). Inflorescence was initiated before current pseudobulb development, which indicated that flowering in Oncidesa was likely controlled by the carbohydrates in back pseudobulbs. Our results showed that there was a noticeable amount of soluble sugars in the current pseudobulb. Still, this only accounted for 27.9% (130 + 130 + 19 mg·g−1) of dry weight of the plants in Expt. I (calculated from Fig. 2A) and 24.1% (830 mg/3.43 g) in Expt. II (calculated from Fig. 5D and Table 3), which was relatively low compared with storage organs of other bulbous crops and the starch in the current pseudobulb was <10% (Figs. 2B, 7A and C). Other carbohydrates in the storage organs of floral crops, such as starch in the bulb of tulip (Tulipa gesneriana ‘Apeldoorn’) was >65% of dry weight (Kamenetsky et al. 2003). Starch was >65% of dry weight in the bulb of lily (Lilium longiflorum Thunb. ‘Nellie White’) (Miller and Langhans 1990) and inulin in the tuber of dahlia (Dahlia spp.) was ∼40% of dry weight (Legnani and Miller 2001; Petkova et al. 2018).
The main carbohydrate in Oncidesa back pseudobulbs was starch, which accounted for >50% of dry weight (Fig. 7A). The starch level in back pseudobulbs of Oncidesa was similar to storage organs in other crops, where the primary storage organs are in Oncidesa Gower Ramsey. However, the current pseudobulb was an essential nutrient source for inflorescence growth. The younger shoots transported more photosynthate to the inflorescence. In comparison, older back shoots transported out less and kept more photosynthates in their pseudobulbs (Yong and Hew 1995a). In this study, the starch level in older back pseudobulbs was higher (Fig. 7A and C). The starch in A pseudobulb was higher than B pseudobulb at the beginning and the starch in A and B pseudobulbs was higher than C pseudobulb during the second growth cycle (Fig. 7A and C). Although older back pseudobulbs stored more starch, the new growth tended to use the starch in the nearest back pseudobulb (the first back pseudobulb). During the first growth cycle, only the starch content in B pseudobulb (the first back pseudobulb) decreased (Fig. 7B), but starch content decreased in both A and B pseudobulbs (the second and first back pseudobulb, respectively) of the plants with two shoots (Fig. 7D). This also indicated that the photosynthesis of the new developing shoot did not meet its growth needs, and thus the reserve carbohydrate in back pseudobulbs was consumed.
Plants demanded more carbohydrates when the biomass or growth rate was higher (Gent and Seginer 2012). Carbohydrate deficiency suppressed the development of plants (Roldán et al. 1999). The starch in back pseudobulbs of Oncidesa with two shoots decreased significantly during the first growth cycle (Fig. 7D). Thus, pseudobulb volume (Fig. 4), fresh weight (Table 2), dry weight (Table 3), and carbohydrate content of the new shoot (Fig. 5) in the plants with two vegetative shoots were lower than plants with one shoot, which kept their starch level stable in the A pseudobulb after the first growth cycle (Fig. 7B). The number of new shoots affected the development of new pseudobulbs. Plants with more shoots developed smaller and lighter pseudobulbs, which was related to nutrient competition between the shoots.
Oncidesa only has one inflorescence on each shoot in most cases. Higher shoot numbers might increase cut-flower number in future harvests, but it also would have a side effect on the development of the current shoot. Although we did not investigate cut flowers, the new shoot with smaller pseudobulb and lower carbohydrates would produce a cut flower with lower quality. It is crucial to maintain a good source-sink relationship to balance harvest and quality; a proper environment for sufficient photosynthesis, the number of back shoots, and the number of new shoots all need to be considered.
Back shoots accumulated photosynthate and supplied carbohydrates to the new growth. When the demand for photosynthate by the developing new shoots exceeds the supply from the back shoots, the starch in back pseudobulbs would decrease to meet the demand. To the contrary, starch increased. In this experiment, starch in back pseudobulbs decreased during the first growth cycle (A and B pseudobulbs) and increased during the second growth cycle (A, B, and C pseudobulbs; Fig. 7). Plants were trimmed and transplanted before the first growth cycle. It probably took some time for the plants to recover and it was winter with low light intensity during the first growth cycle. In addition, there were more back shoots to produce photosynthates in the second growth cycle. The lack of starch in pseudobulb B might also explain why plants did not flower during the first growth cycle.
The photosynthates partitioning to the current pseudobulb of Oncidesa Goldiana fell from 29% to 11% when the inflorescence grew from 5 cm to mature, then it increased to 33% after inflorescence removal (Yong and Hew 1995b). The carbohydrate changes in the pseudobulbs in our study confirmed the change of sink strength mentioned previously. The current pseudobulb accumulated monosaccharides in the pseudobulb when it actively grew and it fell to a low level during inflorescence growth (Fig. 2). Less photosynthate was needed after inflorescence development that led to starch accumulation in the current pseudobulb (Fig. 7).
The daughter bulb of bulbous crops becomes a mother bulb after a cycle of vegetative and reproductive growth, which is similar to Oncidesa, but the main carbohydrate forms of the daughter and mother bulbs in bulbous crops are the same (Ranwala and Miller 2008). Monosaccharides in the current pseudobulb of Oncidesa were soon used after accumulation. However, the daughter bulb of tulip began to develop after flowering and it stored photosynthates from the leaves that emerged from the mother bulb. After the daughter bulb developed, tulips did not grow leaves or flower until it broke dormancy (Ohyama et al. 1988). The annual growth cycle of tulip was longer than the semiannual growth cycles in Oncidesa. Thus, the daughter bulb of tulip stored polysaccharides rather than monosaccharides, which was the case in Oncidesa.
In summary, the major carbohydrates in the developing pseudobulb of Oncidesa were glucose and fructose. After the plant entered the next growth cycle, glucose and fructose were used and the pseudobulb started to store starch as its primary reserve carbohydrate. Although the surface of back pseudobulbs was wrinkled and the volume of the pseudobulbs was smaller than the current pseudobulb because of water loss, carbohydrate concentration in the back pseudobulb was >50%, which was much higher than the 25% of dry weight in the current pseudobulb. The carbohydrate content in a single back pseudobulb was 1.5 to 2.0 times the carbohydrate in the current pseudobulb, and there were also multiple back shoots on the plant. Back pseudobulbs played a crucial role in carbohydrate storage. However, plants used the carbohydrates in the current pseudobulb first. Glucose and fructose in the current pseudobulb were primarily for plant growth, whereas the starch in back pseudobulbs was for storage.
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