聖誕紅簇葉病病原之鑑定與應用

Abstract

聖誕紅 (Euphorbia pulcherrima) 為台灣重要的盆花作物。市售的聖誕紅品種具有簇葉、矮化、側枝多、苞片多等特性，事實上此為感染聖誕紅簇葉病所造成的病徵，因此這個病害反而帶來聖誕紅盆花的正面應用價值。引起聖誕紅簇葉病的病原是植物菌質體 (phytoplasma)，為無法人工培養、無細胞壁、韌皮部侷限之絕對寄生細菌。按照目前植物菌質體的分類依據，感染聖誕紅之植物菌質體類群屬於第三群 (16SrIII) H 亞群為主，少數品種可偵測到 16SrIII 及 16SrI 複合感染。目前仍缺乏更正式而足夠的科學文獻來闡明聖誕紅受不同類群植物菌質體感染造成之性狀差異，這也造成盆花聖誕紅各品種的真實特性因感染植物菌質體而不易釐清，也難精確利用植物菌質體來精準生產特性穩定的聖誕紅盆花。因此本研究先予蒐集目前市售的聖誕紅主流品種，進行 16S rRNA DNA片段序列分析，確認其感染植物菌質體之類群。之後嘗試以抗生素去除不同聖誕紅品種體內之植物菌質體，以瞭解無植物菌質體感染的各品種真正特性。另外嘗試以菟絲子將台灣現有的各類群菌質體接種至上述無感染的聖誕紅，觀察及記錄其病徵的發展，以瞭解不同類群植物菌質體對聖誕紅性狀之影響。PCR 檢測及增幅DNA片段定序結果顯示，所有偵測品種之聖誕紅皆感染 16SrIII 植物菌質體。抗生素試驗結果顯示，聖誕紅扦插前浸泡四環黴素之聖誕紅品種紅絲絨與桃莉，扦插後其植株生長高度顯著大於對照組，且分枝數顯著減少；品種彼得之星浸泡抗生素之處理組與對照組之株高則無顯著差異，此品種對照組僅冒出新芽，未出現明顯分枝。在植物菌質體接種試驗方面，接種日日春葉片黃化病 (Periwinkle leaf yellowing) 植物菌質體 (16SrI) 之各品種聖誕紅，株高顯著低於對照組，分枝數則與對照組無顯著差異。而接種聖誕紅簇葉病 (Poinsettia witches’ broom) 的聖誕紅分枝誘導性植物菌質體 (Poinsettia branch-inducing phytoplasma, 16SrIII) 之各品種聖誕紅，除株高顯著低於對照組外，分枝數也明顯較多。由此可得知臺灣各聖誕紅品種普遍感染 16SrIII 植物菌質體，且此類植物菌質體的感染，對聖誕紅矮株化與良好的分枝性狀有其必要性。另外，聖誕紅分枝誘導性植物菌質體對促進聖誕紅分枝的效果亦較日日春葉片黃化病植物菌質體為佳，因此仍建議使用此類植物菌質體增加聖誕紅分枝性狀。本論文亦針對治療聖誕紅細菌性葉斑病所使用的之保美黴素 (即鏈土黴素) 殺細菌劑進行噴灑測試，發現於品種彼得之星噴灑此藥，對聖誕紅株高與分枝數皆無顯著影響；於品種紅絲絨與桃莉噴灑此藥劑後，株高也無顯著差異。此結果推論只要按建議劑量施用應可安心以此藥防治細菌性葉斑病，不會影響其盆花矮株與分枝特性。

Poinsettia (Euphorbia pulcherrima) is an important ornamental crop in Taiwan. The commercial cultivars of poinsettia are usually free-branching, which is related to poinsettia witches’ broom. The disease therefore brings the big benefit for the industry of poinsettia. The pathogen of poinsettia witches’ broom is the phytoplasma, which is a wall-less phloem-limited bacterium. Based on the taxonomy of phytoplasmas, the predominant type of phytoplasma which infects poinsettias is named Poinsettia branch-inducing phytoplasma, which belongs to 16SrDNA group III. Few cultivars are infected both by 16SrIII and 16SrI. However, the studies associated with the detailed relationships between poinsettias and phytoplasmas are still rare. The poinsettia industry attempts to realize whether the other groups of phytoplasmas can also cause stunting and branching symptoms on poinsettias for their diverse breeding. In this study, commercial poinsettia cultivars were collected and their nucleic acids were extracted for the PCR-based assays and DNA sequencing. The sequences of 16SrDNA amplified fragments were analyzed to identify the groups of phytoplasmas infecting various poinsettia cultivars. The phytoplasma in poinsettia was further eliminated by the tetracycline treatment to understand the natural feature of each poinsettia cultivar without phytoplasma infection. The phytoplasma-free poinsettias were then inoculated with different groups of phytoplasma to investigate the change of phenotype. The results showed that all of the poinsettia cultivars tested were infected by 16SrIII phytoplasma through PCR and sequence analysis. The poinsettia cultivar Red Velveteen and Dulce Rosa treated with tetracycline were significantly taller and the number of branching was significantly fewer than those treated by water for the controls. The height of cultivar Peter Star did not show significance between control and the tetracycline-treated poinsettias, and the control (no treatment with tetracycline) did not show significant branching but some buds appearing. The tetracycline-treated poinsettias (phytoplasma-free) were then inoculated with periwinkle leaf yellowing phytoplasma (16SrI), and the results indicated that they were significantly shorter than the uninoculated poinsettias, but the number of branching did not show significance. On the other hand, the phytoplasma-free poinsettias inoculated by poinsettia branch-inducing phytoplasma (16SrIII) were significantly shorter and their number of branching was significantly larger than the uninoculated poinsettias. The results revealed that most poinsettias cultivars in Taiwan have already infected with poinsettia branch-inducing phytoplasmas, and the infections is very important and necessary for the branching of poinsettias. The poinsettia branch-inducing phytoplasma (16SrIII) can stimulate more branching on poinsettias than the periwinkle leaf yellowing phytoplasma (16SrI) do. The antibiotic (streptomycin + oxytetracycline) is a useful bactericide for the control of bacterial leaf spot of poinsettias according to the previous study. However, the growers worried that the use of streptomycin + oxytetracycline probably also reduce the concentration of phytoplasmas in poinsettias and inhibit their branching. Our trial demonstrated that the height and the number of branching did not show significant difference between antibiotics-treated plants and controls for the cultivar Peter Star. For the cultivars Red Velveteen and Dulce Rosa did not also show significant difference in the height between antibiotics-treated plants and control. This result suggested that the streptomycin + oxytetracycline should be useful and assured for control of bacterial leaf spot of poinsettias without apparent influence of phytoplasma survival.