蝴蝶蘭瓶苗於礦物元素缺乏下之元素動態變化與生理反應

Abstract

蝴蝶蘭 (Phalaenopsis spp.) 為附生蘭，具肥厚之葉片與肉質根，對養分逆境之耐受性強，反應所需時間長，因此研究相對困難。蘭科植物耐營養逆境之機制未明，有一說法為蘭科植物對養分的分配利用效率高，因此可長時間存活於養分逆境下，但相關研究少。為瞭解蝴蝶蘭於養分逆境下之生存機制，本研究以儲存養分較少之蝴蝶蘭瓶苗為材料，給予氮、磷、鉀、鈣、鎂及鐵元素之缺乏逆境後，從生長調查、元素分析、切片解剖及抗氧化系統酵素分析來探討蝴蝶蘭於養分逆境下之反應與體內元素之動態變化。 本研究將蝴蝶蘭 (Phalaenopsis Sogo Yukidian ‘V3’) 中母瓶苗植入氮、磷、鉀、鈣、鎂或鐵元素缺乏的培養基中誘導缺乏徵狀，植入培養基後新生葉片定義為上位葉。對照組培養於以1/2 MS為基礎培養基，其中含有30.0 mM氮、0.62 mM磷、10.1 mM鉀、1.5 mM鈣、0.75 mM鎂與0.1 mM鐵。將基本培養基中的氮、磷、鉀、鈣、鎂及鐵元素分別移除，其他營養元素濃度不變，即為各缺乏處理所使用之培養基。於培養0、1.5、3、4.5及6個月後取樣，探討蝴蝶蘭養分逆境下之生長、體內元素、生理之動態變化。 大白花蝴蝶蘭於缺氮環境下根長增加，且根部分配到較多的氮，相較於上位葉為較強的積貯。缺氮亦使植株總蛋白質含量下降。缺磷植株外觀未見明顯徵狀，下位葉呈紫紅化，且作為磷之主要供源。缺鉀處理外觀雖與對照組類似但少部分植株有側根生長或根尖壞死之情形。除對照組及缺氮處理外，其餘處理植株之鉀元素會優先分配至上位葉。缺鈣處理根部萎縮或壞死，解剖後可見其維管束細胞與皮層細胞萎縮，維管束末端接近根尖處可觀察到細胞死亡。缺鈣植株之上位葉在培養3個月後開始出現水浸狀或黃化掉落之徵狀，嚴重者擴散至植株頂芽或全株黃化死亡。缺鈣處理之葉片及根部之丙二醛 (malondialdehyde，MDA) 濃度較對照組高。缺鎂處理有上位葉黃化之徵狀，根尖形態與缺鈣相似，且植株總抗壞血酸和抗壞血酸含量增加。缺鐵植株初期根長增加，根域附近之培養基顏色變成黃褐色，4.5個月後有上位葉黃化情形。缺鐵處理下位葉鐵濃度由118 µg·g-1減少至38 µg·g-1，鐵含量由7.5 µg減少至3.1 µg；但對照組與其餘缺乏處理之下位葉鐵濃度及含量均呈上升趨勢。鐵為不易移動之元素，在有外源鐵供應之環境中，下位葉亦為鐵元素積貯之一；無外源鐵之缺鐵環境下，下位葉則為上位葉與根部的鐵之供源。缺鐵處理3個月後葉片過氧化氫酶 (catalase, CAT) 活性減少為0.96 unit·g-1，對照組為1.18 unit·g-1，而由於CAT之中心原子為鐵，葉片CAT活性下降應與鐵供應減少有關。與此同時，葉片抗壞血酸過氧化酶 (ascorbate peroxidase, APX) 活性提高至2.53 unit·g-1，對照組為0.90 unit·g-1。 蝴蝶蘭瓶苗體內元素之動態變化為上位葉及根部為兩個主要的積貯互相競爭的結果。在所有處理中，上位葉元素含量隨培養時間增加而增加，下位葉除鈣及鐵元素外，其餘元素濃度隨培養時間增加呈減少趨勢 (下位葉鈣含量持平，鐵含量除缺鐵處理減少外，其餘處理呈增加趨勢)。根部元素濃度及含量隨繼代週期而變化，在同一週期內元素含量隨培養時間而上升。鈣和鐵此二不易移動的元素，蝴蝶蘭植株下位葉之元素含量佔植株總體含量之比例高。在缺乏處理中，無外源元素供應時，植株僅依靠下位葉所貯藏之元素進行生長，因此影響植體對元素的分配與使用。蝴蝶蘭體內之抗氧化酵素系統於本研究中之變化幅度小，原因可能為元素缺乏並非短時間作用之逆境，植株可以慢慢適應，或並未取樣到植體劇烈變化之時期。

Phalaenopsis spp., which is an epiphyte orchid, has thick leaves and fleshy roots with velamen. Phalaenopsis have a high capacity for nutrient storage, hence they are highly tolerant to nutrient stress and thus it takes a long time for them to exhibit deficiency symptoms. Therefore, studying on phalaenopsis mineral nutrition is relatively difficult. The mechanism of tolerance to nutrient stress in orchids remains unknown. One theory is that orchids can utilize nutrients efficiently; therefore, they can survive for a long time under nutrient stress. However, there is little research to support it. to understand the mechanism, we investigated the dynamic changes of growth, nutrient partitioning, tissue anatomy, and antioxidant system responses under nutrient deficiency stresses. In this study, tissue-cultured plantlets from intermediate cultures of Phalaenopsis Sogo Yukidian ‘V3’ were planted into various nutrient deficiency media to introduce nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg), and iron (Fe) deficiency stresses. The newly grown leaves after subculture were defined as upper leaves. Control plants were subcultured in basal medium which mainly contains ½ Murashige and Skoog medium with 30.0 mM N, 0.62 mM P, 10.1 mM K, 1.5 mM Ca, 0.75 mM Mg, and 0.1 mM Fe. Specific nutrient elements were removed from basal media to produce nutrient-deficient media, while concentration of other elements remained the same. Destructive analyses were conducted after 0, 1.5, 3, 4.5, and 6 months. Phalaenopsis Sogo Yukidian ‘V3’ plants have longer roots under N deficiency stress, and as root was a stronger sink, they gained more N. Total protein concentration decreased under N deficiency stress. Lower leaves of P-deficient plantlets acted as a main source of P and turned purple after culture for 6 months. Some K-deficient plantlets developed lateral roots or exhibited root tip necrosis symptom. Except for control and N-deficient plantlets, nutrient-deficient plantlets translocated most of their K into upper leaves. Calcium-deficient plantlets exhibited atrophied or necrosed roots with atrophied cells at the apical meristem. Calcium-deficient plantlets exhibited water soaked or yellowing upper leaves after culture for 3 to 6 months. The symptoms even spread over entire plantlets. In addition, malondialdehyde (MDA) concentration of Ca-deficient plantlets increased. Magnesium-deficient plantlets also exhibited yellowing upper leaves, and the root tip symptom was similar to that of Ca-deficient plantlets. Total ascorbate and ascorbate concentration of Mg-deficient plantlets increased. Root length of Fe-deficient plantlets increased. Iron-deficient plantlets made the medium turn tawny and exhibited yellowing upper leaves after 4.5 months. In lower leaves of Fe-deficient plantlets, Fe concentration decreased from 118 µg·g-1 to 38 µg·g-1 and Fe content decreased from 7.5 µg to 3.1 µg, while Fe concentration and content increased in other treatments. Lower leaves was one of the sinks of Fe when external Fe was existed but acted as the only source while there was no external Fe supplement. Catalase (CAT) activity of leaves in Fe-deficient plantlets decreased to 0.96 unit·g-1 after 3 months, while control group was 1.18 unit·g-1, which may be attributed to that the central atom of CAT is Fe. Meanwhile, ascorbate peroxidase (APX) activity of Fe-deficient plantlets increased to 2.53 unit·g-1, while control was 0.90 unit·g-1. The dynamic changes of nutrients in phalaenopsis were the results of the competition of two sinks, upper leaves and roots. In all treatments, nutrient contents of upper leaves increased with time. Except for Ca and Fe, the nutrient concentration of lower leaves decreased with time, while Ca did not change with time and Fe increased. Nutrient content of roots increased within the same subcultured cycle. Calcium and iron were considered to be less mobile, hence the content of lower leaves accounted for a high proportion in whole plants content. Within nutrient deficiency treatments, which were without specific nutrient supplement, lower leaves acted as the only source to support the growth of plants, and, therefore, the partitioning pattern of nutrient elements in plants was changed. In this study, changes of antioxidant system in phalaenopsis were little. The reason may be that plants can adapt to nutrient deficiency gradually or the sampling times did not meet the severely changing period.