高溫對稻熱病菌致病性及水稻防禦反應之影響

Abstract

近年來全球暖化問題日益嚴重，水稻作為全球重要糧食作物，栽培過程中常受 Pyricularia oryzae 侵害造成稻熱病發生，環境溫度上升在減少水稻產量同時也可能改變 P. oryzae 與水稻之間之交互作用。為瞭解高溫環境對稻熱病之影響，本研究測試 25 株臺灣本土 P .oryzae 對熱的耐受性，發現相較於 26℃，32℃ 及 35℃ 嚴重抑制 P. oryzae 於燕麥瓊脂培養基之菌絲生長 (平均抑制率分別為 59.1% 及 99.8%) 與產孢能力 (平均抑制率分別為 99.3% 及 99.6%)。使用受高溫影響較少的 P. oryzae 12CY-MS1-2 進行人工接種試驗，相較於日/夜溫 28/26℃ 處理，35/33℃ 處理顯著減少麗江新團黑穀 (Lijiangxintuanheigu, LTH)、高雄 145 號 (Kaohsiung No. 145, KH145) 及臺南 11 號之罹病級數，但加劇日本晴 (Nipponbare, NPB) 之病徵；帶有 Pi2、Pi9 及 Ptr 等抗稻熱病基因之 LTH 及 KH145 單基因品系中，Pi9 及 Ptr 於 LTH 遺傳背景時，高溫下罹病級數略為上升，Pi2 則在兩種遺傳背景均維持高度抗性。高溫雖抑制 P. oryzae 12CY-MS1-2 在植株葉片上的產孢 (35/33℃ 處理抑制率為 78-100%) 及壓器形成能力 (35/33℃ 處理抑制率為 26.4-43.7%)，但仍具有二次傳播及侵染之風險。本研究選擇在臺灣具有優良抗性表現之 Pi9，透過 quantitative reverse transcription PCR (RT-qPCR) 進行基因表現量測定，結果顯示 35/33℃ 高溫環境下 Pi9 仍能表現，與其對應之稻熱病菌 avirulence (Avr) 基因 AvrPi9 於部分品種則表現量提升，顯示高溫強化 P. oryzae 侵染水稻之能力。進一步以 RT-qPCR 檢測活性氧生成基因 respiratory burst oxidase homologues B (OsRbohB) 及受植物賀爾蒙水楊酸 (salicylic acid, SA) 與茉莉酸 (jasmonic acid, JA) 調控之稻熱病抗性相關基因之表現量，發現不同品種應對高溫逆境之調控方式有所差異，但部分基因對高溫處理呈現相同趨勢，如高溫抑制 OsRbohB、SA 訊號傳遞基因 nonexpresser of pathogenesis-related genes 1 (OsNPR1) 及 JA 生合成基因 allene oxide synthase 2 (OsAOS2) 之表現；而 SA 訊號傳遞基因 WRKY DNA-binding protein 45 (OsWRKY45) 與 JA 訊號傳遞基因 jasmonate-zim-domain protein 9 (OsJAZ9) 於高溫下受到誘導；防禦下游基因 pathogenesis related protein 1b (OsPR1b) 則在 LTH、KH145 及 NPB 於侵染早期受高溫抑制；與非生物及生物逆境相關基因 stress associated protein 1 (OsSAP1) 則在高溫下受到誘導，並於 KH145 遺傳背景中有更高的表現，顯示 KH145 可能具有較好的耐非生物或生物逆境之能力。帶有 Pi9、遺傳背景為 KH145 之水稻品種臺大高雄 1 號 (National Taiwan University-Kaohsiung No. 1, NTU-KH1) 於高溫環境下維持對稻熱病高度抗性，且多種參與非生物逆境之基因 (OsRbohB、OsWRKY45 及 OsJAZ9) 於高溫環境上調，顯示具有抗非生物逆境之潛力。本研究探討高溫下水稻與 P. oryzae 之間的交互作用，期望為稻作病害管理提供科學依據，以確保糧食安全，減少氣候變遷帶來的潛在損失。

In recent years, the problem of global warming has become increasingly serious. Rice, as one of the world’s most important staple crops, is often attacked by Pyricularia oryzae, which causes rice blast during cultivation. Rising temperatures not only reduce rice yields, but may also alter the interaction between P. oryzae and rice. To understand the impact of high temperature on rice blast, this study tested the heat tolerance of 25 P. oryzae isolates from Taiwan. Compared to 26℃, both 32℃ and 35℃ significantly inhibited the mycelial growth (average inhibition rates were 59.1% and 99.8%, respectively) and sporulation ability (average inhibition rates were 99.3% and 99.6%, respectively) of P. oryzae on oatmeal agar medium (OMA). The inoculation experiments were conducted using P. oryzae 12CY-MS1-2, which was less affected by high temperatures. Compared to the treatment with day/night temperature of 28/26℃, the 35/33℃ treatment significantly reduced the disease severity in rice varieties Lijiangxintuanheigu (LTH), Kaohsiung No. 145 (KH145), and Tainan No. 11, but enhanced the disease symptoms in Nipponbare (NPB). Among the LTH and KH145 monogenic lines carrying blast resistance genes Pi2, Pi9, and Ptr, Pi9 and Ptr showed a slight increase in disease severity under high temperatures in the LTH genetic background, while Pi2 maintained high resistance in both genetic backgrounds. Although high temperatures suppressed the sporulation of P. oryzae 12CY-MS1-2 on rice leaves (inhibition rates were 78-100% at 35/33℃) and reduced its appressorium formation ability (inhibition rates were 26.4-43.7%), there remains a risk of secondary transmission and infection. This study selected Pi9, which has been highly resistant in Taiwan, and quantified its expression through quantitative reverse transcription PCR (RT-qPCR). The results showed that Pi9 remained expressed in the high temperature environment of 35/33℃. The corresponding avirulence (Avr) gene of P. oryzae, AvrPi9, exhibited increased expression in some varieties under high temperature, suggesting that elevated temperature enhances the virulence of P. oryzae. RT-qPCR was also conducted to quantify the expression of reactive oxygen species (ROS)-producing gene respiratory burst oxidase homologues B (OsRbohB) and the blast resistance-related genes regulated by plant hormones salicylic acid (SA) and jasmonic acid (JA). The results revealed differences in regulatory responses to high temperature among different rice varieties. However, some genes showed similar trends in response to high temperature treatment. For example, high temperature suppressed the expression of OsRbohB, SA signaling gene nonexpresser of pathogenesis-related genes 1 (OsNPR1), and JA biosynthesis gene allene oxide synthase 2 (OsAOS2). Conversely, SA signaling gene WRKY DNA-binding protein 45 (OsWRKY45) and JA signaling gene jasmonate-zim-domain protein 9 (OsJAZ9) were induced under high temperature. The downstream defense gene pathogenesis related protein 1b (OsPR1b) was suppressed in LTH, KH145 and NPB during the early stage of infection under high temperature. Stress associated protein 1 (OsSAP1), a gene related to abiotic and biotic stresses, was induced under high temperature and showed higher expression in KH145 genetic background. This indicates that KH145 may have better tolerance to both abiotic and biotic stresses. The rice variety National Taiwan University-Kaohsiung No. 1 (NTU-KH1), which carries Pi9 in the KH145 genetic background, maintained high resistance to rice blast under high temperature, and a variety of genes involved in abiotic stress (OsRbohB, OsWRKY45 and OsJAZ9) were upregulated under high temperature, indicating the potential to resist abiotic stresses. This study investigated the interaction between rice and P. oryzae under high-temperature conditions, offering a scientific basis for rice disease management to enhance food security and reduce potential losses driven by climate change.