(64(2):89-98)Effect of Temperature on Virulence of Ralstonia solanacearum Biovars and Response of Potato Cultivars (Lines) to Bacterial Wilt

青枯病菌 (Ralstonia solanacearum) 引起之細菌性萎凋病為熱帶及亞熱帶地區作物關鍵病害之一。多數青枯病菌株於高溫對寄主具有病原性，部分菌株在20℃以下則會失去其病原性。然而，分類上屬於第三生理小種、第二生化型 (R3Bv2) 的菌株於冷涼的溫度條件下仍可感染寄主植物，造成病害。本研究中調查2013 年11月至2014年3月期間，台灣雲林縣及嘉義縣馬鈴薯栽培區萎凋病之發病率為0–32%，分離其病原菌，發現皆為青枯病菌，其中64.7% 之分離株為第二生化型之菌株，其餘為第三生化型之菌株，顯示青枯病菌第二生化型之菌株為目前危害台灣馬鈴薯之主要菌株。此外，亦於不同溫度下比較台灣常見3 種R. solanacearum生化型菌株對茄科作物之致病力差異。結果顯示在24℃及28℃，3 種生化型菌株 (RS1102、RS1118 及RS1125)皆對試驗植物具有良好的致病力，接種後20 d 罹病度 (disease severity) 介於77–100%。但於20℃時，RS1125(第三生化型) 則對馬鈴薯失去其病原性，而RS1118 (第二生化型) 則表現較其他兩種生化型菌株較強之毒力，接種20 d 後馬鈴薯及番茄上之罹病度為80–100%。青枯病為馬鈴薯生產栽培之限制因子之一，故本研究利用10 種不同之馬鈴薯品種 (系) [P86 (「克尼伯」)、P90 (「大西洋」)、P91 (「種苗2 號」)、P92 (「吉比 達」)、P95 (‘pike’)、P117 (‘Chipeta’，「聯華」)、P120 (「台農1 號」)，P89、16-13 及T3] 針對第二生化型之菌株進行抗病分析，以期篩選對青枯病具有抗性之品種。結果顯示，品種間表現出不同程度之耐性，但未獲得表現高抗 (無萎凋) 之抗病品種。接種35 d 後，品種間罹病度為10–100%，其中以T3 品種之罹病度最低。經上述試驗結果可知，溫度為影響青枯病發病率之因素之一，台灣常見之3 種青枯病菌生化型菌株中，則以第二生化型菌株在低溫時對馬鈴薯之危害程度為最高、威脅性最大。 Bacterial wilt, caused by Ralstonia solanacearum, is an important disease of a wide range of crops in tropical and subtropical regions. Most strains of R. solanacearum are pathogenic at high temperatures, and some strains are nonpathogenic when the temperature drops below 20℃. However, race 3 biovar 2 (R3Bv2) strains can attack plants at cool temperatures. In this study, we surveyed the potato fields in Yunlin and Chiayi Counties in Taiwan during 2013–2014 and observed the incidences of wilting symptoms on potato, within 0–32%. The causal pathogens were identified and most of the stains were identified to be R. solanacearum biovar 2, indicating that the biovar 2 is a dominant strain to affect potato in Taiwan. In addition, the effects of temperature on the virulence of three R. solanacearum strains (RS1102, RS1118 and RS1125) and the representative biovars affecting solanaceous crops in Taiwan were evaluated on potato and tomato plants, respectively. Results indicated that all three strains showed strong virulence on potato and tomato plants at 24℃ and 28℃, and the disease severity ranged from 77 to 100% at 20 days of post inoculation. At 20℃, the three strains showed differential reactions on the inoculated potato. Among them, RS1125 (biovar 3) was nonpathogenic to potato, whereas RS1118 (biovar 2) showed higher virulence than the other two strains with 80–100% disease severity at 20 days of post inoculation. To compare the resistance between the current commercial potato cultivars to R. solanacearum race 3 biovar 2, ten cultivars [P86 (‘Kennebec’), P90 (‘Atlantic’), P91 (‘Tss-II’), P92 (‘Chipeta’), P95 (‘pike’), P117, P120 (‘TN-1’), P89, 16-13, and T3] were examined in this study. It indicated a differential susceptibilities among them, from susceptible to various degrees of tolerance, to the infection of R. solanacearum race 3 biovar 2. The disease severity ranged between 10–100% at 35 days of post inoculation. One cultivar, T3, showed the lowest disease severity at 10%. No cultivar exhibited immunity to the bacterium. Based on our results, we concluded that temperature is one of the factors that can affect the virulence of R. solanacearum. Comparing the virulence of three commonly found biovars of R. solanacearum in Taiwan at different temperatures, we suggest that bovar 2, which showed high virulence at cool temperatures, might be a great threat to the potato production in Taiwan

Bacterial soft rot of radish in Taiwan

自臺灣中西部不同地區之蘿蔔田，共分離Erwinia 屬軟腐細菌５０個菌株，依其生理 生化特性測定結果，其中４６個菌株為 E．carotovora subsp．carotovora，而其餘 ４個菌株則為E．chrysanthemi，此４個 Ech菌株均屬於Dickey氏區分 Ech之subdivi sionⅡ．分離之菌株中未發現E．carotovora subsp．atroseptica．Ecc與Ech在十種 蔬菜組織均可引起軟腐，但Ech 之致腐能力常較Ecc 為強，尤其在洋蔥鱗片上之軟腐 能力差異更大．以三種品種洋蔥作接種測定，結果大部份蘿蔔之Ecc 菌株，在洋蔥鱗 片上之軟腐能力很弱，而蘿蔔之Ech 菌株則可造成較大之軟腐. 溫度影響蘿蔔軟腐菌株之致腐能力，但２４℃以上之溫度，Ech 致腐能力常高Ecc ． 以等量之Ecc與Ech同時接種蘿蔔，在較高之溫度（３２℃），軟腐之蘿蔔組織中以Ec h 佔優勢，於nutrient broth測定其生長速率，在３２℃以上，Ech 生長較Ecc 為快 ．氧氣亦可影響軟腐細菌之致腐能力，無氧狀態，僅需較低之菌量（１０^2－１０^3 cells），即可造成５０％之軟腐發病率，有氧時則需較高之菌量（１０^4 cells） ．不同部位軟腐之程度亦有差異，而以蘿蔔上半部位軟腐較小． 比較各種土壤添加物對蘿蔔細菌性軟腐病發生之影響，施用SH混合物或烏肥與當地農 民使用之添加物（含稻穀、尿素），在自然與人為接種之發病率，並無差異，施用SH 混合物與烏肥者，其總產量增加，貯藏根亦較大．自田間取回蘿蔔貯存根，於實驗室 進行接種試驗，結果以施用有機肥者腐爛最小，而SH混合物者次之，施用烏肥者，則 較其它處理為感病．於實驗室測定不同添加物對軟腐細菌存活之影響，SH混合物、烏 肥或未處理之土壤，其族群均很快下降，惟處理烏肥者下降較快．於蘿蔔田施用SH混 合物，在作物生長後期，土中軟腐細菌族群仍可達１０^3－１０^5 cfu／g土壤．溫 室試驗結果，軟腐細菌在蘿蔔根圈土壤，可維持較長時間

Erwinia carotovora subsp. carotovora與Erwinia chrysanthemi引起籮蔔細菌性軟腐病

Bacterial soft rot of radish frequently occurs in Taiwan and was reported to be caused by Erwinia carotovora subsp. carotovora (Ecc). In 1988, a total of fifty strains of soft-rotting Erwinia was isolated from diseased radish tissues collected from several fields in the central and southern parts of Taiwan, among them, forty-six strains were identified as Ecc, while the other four strains were Erwinia chrysantherni (Ech) with phenotypic characteristics similar to the strains in subdivision II according to Dickey’s classification. Strains of both Ecc and Ech induced soft rot extensively on a wide variety of vegetable tissues. However, Ech strains were more virulent to bulb scales of onion than most Ecc strains. The rotting ability of Ech on radish taproot tissues was also stronger than that of Ecc especially at higher temperature (32℃). In addition1, when radish taproot tissues were inoculated with cell suspensions of both Ecc and Ech in a 1:1 ratio, Ecc was dominant in the rotted tissues at 28℃ or below, but Ech became dominant at higher1temperature (32℃). 蘿蔔細菌性軟腐病於台灣田間經常可見，然對其病原細菌之特性，未曾有較深入之探討。民國77年間筆者等自台灣中南部不同地區之蘿蔔田，採回罹軟腐病之蘿蔔塊根與葉柄組織，共分離了50個Erwinia屬軟腐細菌菌株，經生理生化特性測定結果，其中46個菌株為Erwinia carotouora subsp. carotouora (Ecc)，另4個菌株為Erwinia chrysanthemi (Ech)，此4個Ech菌株皆屬於Dickey分類體系之第II型。Ecc與Ech菌株對許多蔬菜組織常具有強之致腐能力，然而Ecc與Ech菌株在洋蔥鱗片上之致腐能力，則有很大差異，許多Ecc菌株對洋蔥鱗片之致腐能力很弱，而Ech菌株則很強。又溫度可影饗此兩種軟腐細菌之致腐能力，於蘿蔔塊根組織上，Ech菌株常較Ecc菌株之致腐能為強，尤其於較高的溫度下(32℃)。以等量之Ecc與Ech細菌同時接種於蘿蔔塊根組織，在較高的溫度下(32℃)，於軟腐組織中Ech之族軍較Ecc為優勢

(48(4):64-70)Effects of Peat Moss Amendment on Propagation of Arbuscular Mycorrhizal Fungi

叢枝菌根菌Glomus mosseae 的繁殖一般採用河砂為栽培介質，是因河砂具有高透氣性與低肥份的特性，但缺點是保水性差及不具緩衝性，泥炭苔則具有高保水性的優點，因此本試驗目的是探討河砂中添加泥炭苔，提高叢枝菌根菌在宿主根系的繁殖效果。試驗首先將泥炭苔（10%）添加方式分成混合或覆蓋兩種處理，供試宿主植物有百喜草、營多藤、玉米及甘藷等四種，結果除甘藷外，菌根形成率與孢子濃度都以混合處理者高於覆蓋處理者。將河砂混合1/5, 1/3, 1/2（體積／體積）的泥炭苔，或單獨使用河砂，探討對G. mosseae 在百喜草根圈土繁殖之影響，結果顯示盆缽中完全為河砂者，菌根形成率與厚膜孢子數最高，隨著泥炭苔混合比例增加，菌根形成率與厚膜孢子數逐漸減低。泥炭苔採用覆蓋方式，定期調查 G. mosseae 在玉米根圈土的繁殖，顯示隨接種時間增加，菌根形成率與孢子數逐漸提高，至接種後第11週達到高峰，而後逐漸降低。由以上結果，顯示單獨以河砂為介質，所生產的孢子數最高，添加泥炭苔反而不利於菌根形成與孢子的產生，其可能的原因有待進一步探討。Sand is a good medium for inoculum production of vesicular-arbuscular mycorrhizal fungi, because of its high aeration and lower fertility, but it has poor water-conserved and buffering ability. In contrast, peat moss has high water-conserved ability, so we try to amend sand with peat moss to improve initial growth of host plants, and to increase colonization and spore of VAM fungi. Effect of 10% peat moss mulched or mixed with sand on colonization and spore production of G. mosseae are tested in four host plants of bahiagrass, intortum, corn, and sweet potato. Results indicated sand mixed with peat moss is better than those mulched with peat moss. Then, the effects of sand mixed with peat moss in ratio of 1/5, 1/3, 1/2 on propagation of G. mosseae in rhizospheric soil of bahiagrass were conducted. Results indicated that the amendment of peat moss in sand could improve initial growth of host plants, but colonization or spore production was the highest in treatment without peat moss amendment. When amended volume of peat moss was over 1/4, there was no spore in rhizospheric soil at 14 weeks after inoculation. Periodic investigation were conducted to monitor propagation of G. mosseae in the rhizosphere of corn, and results indicated that root colonization and spore production in soil gradually increased, and reached the highest at 11 weeks after inoculation, and then decreased. The results showed that spore production was the highest in sand without any amendment. Peat moss was not beneficial to colonization and spore production, if amended with sand

Development of Mass Production of VAMF Inoculum by Sand Culture

內生菌根菌(Vesicular-aibuscular mycorrhizal fungus, VAMF)證實與許多作物根系具有親和主，當VAMF感染作物根部形成內生菌根時，往往能產生有益效應，促進根群發育，增加根部對磷肥等要素之吸收能力，促進植株生長。惟內生菌根菌是一種活物寄生真菌，目前尚無法在文工培養基上大量繁殖，接種盆栽宿主植物繁殖內生菌根菌接種源是目前唯一可行途徑。除宿E外，土壤（栽培介質）因子、環境因子等亦可影響內生菌根菌之繁殖。本分所建立之內生菌根百接種源生產體系，分為三個步驟：（一）以多年生百喜草盆栽保存純種，（二）以盆栽玉米繁殖貝種，（三）以大型栽培床之玉米（或營多藤）大量繁殖內生菌根菌，經收集培養土（含宿主根長）風乾裝袋而成接種源。在實際應用上，採用苗期接種，培養洋香瓜內生菌根苗，當內生菌根苗移植於本田，即可將菌種帶到田間，發揮內生菌根之功能，表現其有益效應。 The vesicular-arbuscular mycorrhizal fungus was proved to have the compatibility with root systems of many crops. Formation of VA mycorrhiza in root systems of crops infected by VAM fungus could arise beneficial effects, i.e. promoted the growth and development of root systems, increased the absorption of soil mineral elements, enhanced the top growth of crops. Up today, the common method for propagation of VA mycorrhizal fungi were pot culture because they were obligate symbionts. Factors affected the propagation of VA mycorrhizal fungi in pot culture included host plants, soil (culture media), and environment. We developed a procedure for propagation of VAM fungus inoculum as follows: 1. maintaining the pure VA mycorrhizal fungus in pot culture of bahiagrass, 2. propagation of VAM fungus in pot culture of corn, 3. mass production of VA mycorrhizal fungus inoculum in sand bed culture of corn or beggarhce, 4. air-drying and bagging the harvested culture soil including the segments of infected root systems of host plants and fungal chiamydospores. Method for application of VA mycorrhizal fungus to field was by transplanting mycorrhizal seedlings of muskmelon pre-inoculated with infested soil in plastic net house. When the mycorrhizal seedlings were transplanted to the field, mycorrhizal root systems could show beneficial effects to muskmelon production

ccurrence and integrated pest management of plant parasitic nematodes in Taiwan

本省植物病原線蟲經調查計有31 屬41 種寄生性線蟲為害本省經濟作物而其中導致本省作物連作障害的線蟲，首推根瘤線蟲（Meloidogyne incognita, M. javanica, M. arenaria, M. hapla ) ，其次為南方根腐線蟲（Pratylenchus coffae）及柑桔線蟲（Tylenchulus semiPenetrans）。此外，地上部植物病原線蟲如葉芽線蟲( APhelenchoides besseyi ）及松材線蟲（BursaPhelenchus xylophilus ）在本省水稻及二葉松等作物上亦造成重大危害。本省作物線蟲病害之防治宜採用健康種苗，選植抗線蟲之品種或砧木，配合非寄主作物輪作制度、土壤有機質添加、生物防治、太陽能土壤消毒及化學葯劑以消滅或管理降低土壤中病原線蟲之密度，以減少線蟲之危害，並達成永續管理之目地。 There are 31 genera and 41 species of plant-parasitic nematodes recognized in Taiwan, among them, root-knot nematode (Meloidogyne incognita, M. javanica, M. arenaria, M. hapla), root-lesion nematode (Pratylenchus coffeae) and citrus nematode (Tylenchulus semipenetrans) were the major found in continuously cropping problem due to phytonematodes. Management of plant parasitic nematodes problem in this report rely on specific basic strategies to kill and decrease the inoculum density of pathogen, including using healthy seedling / resistant stock, non-host rotation, adding organic amendments, biological control, urea treated with solarization and/or chemical control

Occurrence of Foot Rot Disease of Sweet Potato Caused by Phomopsis destruens in Taiwan

甘藷基腐病於2008 ~ 2011 年間於台灣之甘藷田陸續發現，受害植株地上部生長勢衰落、葉片黃化枯萎、藤蔓乾枯死亡，由於乾枯部位緊鄰塊根生長處，因此導致罹病植株幾乎完全無法生產塊根，且由本病害亦會感染塊根組織部位，造成水浸狀影響甘藷品質。本研究自罹病病株莖蔓分離得到菌株(SPPD-1)，以PDA 培養時，該菌菌絲生長緩慢，於25℃ 下培養20 天後，培養基呈現淡褐色，菌落稍有皺摺、邊緣不整形。本菌會產生兩型分生孢子，其中一型為單胞、透明無色、圓筒狀或卵形、兩端具有圓形油滴之甲型分生孢子(α-conidia)，於罹病組織或人工培養過程中皆會產生；另一型則為次紡錘型，具多細油滴、一端突尖或圓鈍，另端稍呈截頭狀，一邊略彎之丙型分生孢子( γ-conidia)，僅偶爾於罹病組織上發現，以PDA 培養時並未產生絲狀或一端彎曲之乙型分生孢子。依據本病害之病徵表現、病原菌形態特徵、分子標記及文獻資料比對，將此菌鑑定為Phomopsis destruens (Harter) Boerema。經人工接種孢子懸浮液於台農57 號甘藷健康苗後，發現P. destruens 對甘藷具有病原性，為引起甘藷基腐病之病原菌。本病害描述及病原記錄為台灣病害之首報。 Foot rot disease of sweet potato (Ipomoea batatas) was found in major sweet potato production areas in Taiwan during 2008-2011. The symptoms include yellowing of leaves, necrotic lesions on vine, wilting of the sweet potato plants. The fungus isolated from diseased tissue grew slowly. The fungal colony could produce brown pigments and diffused into the agar medium cultured on PDA at 25℃ for 20 days, and also displayed slightly zonate, sulcate and wavy edge. The fungus generated two types of conidia from stromatic pycnidia. One type conidia were one-celled, hyaline, oblong or oval, 2-guttulated α-conidia；the other type conidia were clavate subcylindrical slightly curved γ-conidia only found on diseased host tissues. The causative fungus was identified as Phomopsis destruens (Harter) Boerema based on symptoms, morphological characteristics and ITS rDNA sequence blasting on NCBI GenBank database. The pathogenicity test of P. destruens toward sweet potato host was proved and fulfilled the Koch’s postulates. To our knowledge, this is the first report on the occurrence of foot rot disease of sweet potato caused by Phomopsis destruens in Taiwan

Surveillance and Integrated Diseases and Pests Management of Ornamental Plants

本計畫調查景觀作物主要病蟲害種類及其週年消長，全年監測有無列名檢疫有害生物之發生情形，並測試主要病蟲害之防治藥劑效果並研發主要的非農藥防治技術。建立重要景觀作物有害生物相，建立防治病蟲害之環境管理規範及非疫生產點技術之研發。 This project focus on the study of management of insect pests and diseases of Ornamental plantsall year around. Some quarantine pests listed were surveyed and monitored. Development of the recommended measures and schedules and nonchemical control of pests and diseases were urgent requirement. The standard operation processes for pests and diseases control will be built up