摘要
水稻生长发育过程中需要应对生物及非生物胁迫。可以通过水稻种子传播的稻瘟病(Magnaporthe oryzae)和恶苗病(Gibberella fuiikuroi)是危害水稻生产的重要真菌病害,白叶枯病(Xanthomonas oryzae pv.oryzae)和细菌性条斑病(Xanthomonas oryzae pv. oryzicola)是重要的细菌病害。土壤中累积的多环芳香烃物质是影响水稻种子萌发、秧苗生长和产量的化学胁迫因子之一。防控水稻种子传播的重大病害、挖掘降解多环芳香烃物质的微生物资源对水稻安全生产具有重要意义。本研究据此开展了水稻种子处理药剂混配研究以其为防控水稻种传病害提供技术支持；分离自黑龙江水稻种子的菌株HLJ-RS18为多环芳香烃的降解提供了新的生物修复菌株,以及对三环芳香烃菲的降解途径展开研究。相关研究结果如下：
1)防治水稻种传真菌病害和细菌病害的杀菌剂混配研究：以防治真菌病害的杀菌剂戊唑醇、咪鲜胺和防治细菌病害的杀菌剂噻菌铜为有效成分,通过菌丝生长速率法、混浊度法等进行毒力测定和联合毒力评价,表明杀菌剂咪鲜胺对水稻稻瘟病菌和恶苗病菌具有显著抑制作用,其ECso分别为0.06μg/ml和0.04μg/ml;戊唑醇对两种真菌的EC5o分别为2.43μg/ml和0.10μg/ml。噻菌铜对白叶枯病病、细菌性条斑病的ECso分别为30.20μg/ml和61.06μg/ml.混配药剂室内毒力测定结果显示,戊唑醇和噻菌铜按照9：1、咪鲜胺和噻菌铜按照7：3混配,对稻瘟病菌和恶苗病菌具有加和或者增效作用,同时对白叶枯病病菌、细菌性条斑病菌亦具有增效或加和作用。在此基础上,确立了适合于水稻种传病原真菌和细菌消毒处理的混配制剂—戊唑醇和噻菌铜混配制剂、咪鲜胺和噻菌铜混配制剂,获得2项中国发明专利,为防控水稻种传真菌和细菌病害的传播蔓延和发生危害提供了新的技术支持。
2)水稻种子寄藏新鞘氨醇杆菌属细菌HLJ-RS183(?)种鉴定及其对芴的降解：从来自于黑龙江的水稻种子样品中分离得到]HLJ-RS18,为Novosphingobium属分离物；经16S rRNA,系统发育树、表型特征、生理生化、化学分类特征等数据分析表明,HLJ-RS18菌株为新鞘氨醇杆菌属的一个新种,命名为Novosphingobium fluoreni sp. nov. HLJ-RS18T。该菌已经获得国际菌种保藏机构的认定,菌株备存于德国DSMZ (=DSM27568T)和中国农业微生物菌种保藏管理中心(=ACCC19180T)。 HLJ-RS18能够降解三环芳香烃芴,降解常数为0.13d-1,半衰期为2.76d。HLJ-RS18对四种供试水稻品种的发芽率和发芽势无影响,对稻瘟病菌和恶苗病、白叶枯病病和细菌性条斑病的供试菌株的生长无影响；分析认为,]HLJ-RS18对多环芳香烃物质的降解具有潜在的应用价值。
3) Stenotrophomonas maltophilia C6菌株降解菲的研究：研究结果显示,S.maltophilia C6能够在14d内完全降解50ug/ml的菲；从C6菌株和菲共培养液中分离鉴定了22个代谢中间产物,通过对代谢产物的分析,发(?)S. maltophilia C6菌株能够通过1,2-,3,4-,及9,10-C途径降解菲,进一步对1,2-,3,4-,及9,10-C途径上层代谢物的总浓度分析,表明菌(?)S. maltophilia C6对菲的降解主要通过3,4-C途径,这是首次对S. maltophilia降解菲的代谢途径的报道。S. maltophilia C6对多环芳香烃物质的降解具有潜在的应用价值。
Plants suffer from both biotic and abiotic stress. Magnaporthe oryzae, Gibberella fujikuroi, Xanthomonas oryzae pv.oryzae, and Xanthomonas oryzae pv. oryzicola are the main rice pathogen that cause fungi and bacteria disease, which bring enormouse economic losses. Polycyclic Aromatic Hydrocarbons (PAHs), the most prevalent and persistent pollutants in the environment, could affect the germination and growth of plants and verified in many references. It is vital to control rice main fungi and bacteria disease and put efforts on bioremediation microbes. This research forcus on both the rice seed chemicals treatment to protect rice from pathogen affected and studies on bioremediation microbes to get possibilities to remove PAHs pollutions. The main results are as below:
1) Seed treatment with chemicals mixture research. Tebuconazole and prochloraz are chemicals that registered as fungicide, and thiediazole copper were used to kill bacteria. To well protect seeds from both fungi and bacteria disease, a mixture proportion of tebuconazole and thiediazole copper or prochloraz and thiediazole copper to prevent rice from seed-borne bacteria and fungi disease were conducted. Tebuconazole and thiediazole cooper mixed as9:1is the optimum proportion as seed treatment, which could well protect rice seeds from both fungi and bacteria. Also prochloraz and thiediazole copper mixed as7:3could protect rice seeds from both fungi and bacteria infection. Two patents were granted by the China patent office for the two chemicals mixture used for seed treatment, which gave techonoly supports for controlling rice seed-borne bacteria and fungi disease.
2) Identification and classification of fluorene-degradation species Novosphingobium fluoreni sp. nov. HLJ-RS18T.We isolated a bacteria strain HLJ-RS18from Sanjiang rice variety of Heilongjiang Province of China, which could degrade fluorene (three rings of polycyclic aromatic hydrocarbons/PAH). Further analysis of16S rRNA, genotypic characterization, chemotaxonomic results and phenotypic analysis indicated that HLJ-RS18T represents a novel species in the genus Novosphingobium. Therefore, we propose the species Novosphingobium fluoreni sp. nov. with HLJ-RS18T (=DSM27568=ACCC19180) as the type strain. HLJ-RS18T showed no effect on the germination of four rice varieties provided. There was no interaction of strain HLJ-RS181with Magnaporthe oryzae YN08-1, Gibberella fujikuroi US01, Xanthomonas oryzae pv.oryzae PXO99, and Xanthomonas oryzae pv. oryzicola RS105.HLJ-RS18T is a new potential bioremediation strain that could be used in the PAHs degradation.
3) Multiple degradation pathways of phenanthrene by Stenotrophomonas maltophilia strain C6. Stenotrophomonas maltophilia C6was isolated by our cooperation lab, and strain C6could completely degrade phenanthrene through14days. To further well understanding the phenanthrene pathway, we isolated and identified22metabolites from the culture periodically. Finally we proposed the phenanthrene pathway by strain Stenotrophomonas maltophilia C6. Stenotrophomonas maltophilia C6could degrade phenanthrene starting from1,2-,3,4-,9,10-C position and following with ortho and meta cleavage. The concentration analysis of upper metabolites from1,2-,3,4-,9,10-C pathway, respectively, showed strain Stenotrophomonas maltophilia C6degrade phenanthrene mainly through3,4-C pathway. This is the first study of detailed phenanthrene metabolic pathways by Stenotrophomonas maltophilia. Stenotrophomonas maltophilia C6is also a potential biomediation bacteria that could contribute to the remove of phenanthrene with clearly proposed degradation pathway.
引文
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