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Echinochloa Crus-Galli This article was downloaded by: [Jianping Zhang] On: 19 May 2014, At: 18:36 Publisher: Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK Biocontrol Science and Technology Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/cbst20 Bipolaris eleusines, a potential mycoherbicide candidate for control of barnyardgrass (Echinochloa crus-galli) Jianping Zhanga, Gary Pengb, Guifang Duana, Yongjun Zhoua, Shuang Yanga & Liuqing Yua a State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, China b Agriculture and Agri-Food Canada, Saskatoon, SK, Canada Accepted author version posted online: 10 Apr 2014.Published online: 14 May 2014. To cite this article: Jianping Zhang, Gary Peng, Guifang Duan, Yongjun Zhou, Shuang Yang & Liuqing Yu (2014) Bipolaris eleusines, a potential mycoherbicide candidate for control of barnyardgrass (Echinochloa crus-galli), Biocontrol Science and Technology, 24:7, 839-846, DOI: 10.1080/09583157.2014.891724 To link to this article: http://dx.doi.org/10.1080/09583157.2014.891724 PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms- and-conditions Downloaded by [Jianping Zhang] at 18:36 19 May 2014 Biocontrol Science and Technology, 2014 Vol. 24, No. 7, 839–846, http://dx.doi.org/10.1080/09583157.2014.891724 SHORT COMMUNICATION Bipolaris eleusines, a potential mycoherbicide candidate for control of barnyardgrass (Echinochloa crus-galli) Jianping Zhanga, Gary Pengb, Guifang Duana, Yongjun Zhoua, Shuang Yanga and Liuqing Yua* aState Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, China; bAgriculture and Agri-Food Canada, Saskatoon, SK, Canada (Received 5 November 2013; returned 9 January 2014; accepted 3 February 2014) An isolate of Bipolaris eleusines was investigated as a potential candidate for biocontrol of barnyardgrass and additional weeds in paddy rice fields and for safety to selected crop species under greenhouse conditions. Barnyardgrass appeared more susceptible at the three-leaf stage than older plants, and disease severity increased as the fungal inoculum increased from 1 × 105 to 1 × 107 conidia/ml when sprayed till run-off. The high application rate caused 73% mortality of barnyardgrass, relative to the non-treated control, but increasing application rate to >1 × 107 conidia/ml did not enhance efficacy (P>.05). This B. eleusines isolate showed no pathogenicity to rice (Oryza sativa spp. indica, O. sativa spp. japonica and an O. sativa hybrid), corn, wheat or any dicot crop species tested while causing slight infection on sorghum and barley. We conclude that B. eleusines, with high efficacy against barnyardgrass and demonstrated safety to rice, is a promising mycoherbicide candidate worthy of further evaluation and development for control of barnyardgrass in paddy rice fields. Keywords: conidia; weed fungal pathogen; Oryza sativa; rDNA; ITS Barnyardgrass [Echinochloa crus-galli (L.) Beauv], the worst weed in paddy rice, is an annual species native to Asia but can now be found worldwide (Holm, 1977). Infestations can reduce rice yields by 44–96% (Ampong-Nyarko & De Datta, 1991; Islam & Kato-Noguchi, 2013). Repeated use of herbicides has resulted in herbicide- Downloaded by [Jianping Zhang] at 18:36 19 May 2014 resistant biotypes of barnyardgrass (Vidotto, Tesio, Tabacchi, & Ferrero, 2007; Wu, 2007). Additional strategies that alleviate the reliance on chemical herbicides are needed for their sustainable management. Several species of Helminthosporium sensu lato, including Pyrenophora graminea; Cochliobolus lunatus; Drechslera monoceras and Exserohilum monoceras, have been reported for potential use against barnyardgrass or other weeds (Kadir, Sajili, Juraimi, & Napis, 2008; Zhang & Watson, 1997; Zhang, Yang, Zhou, & Yu, 2010). However, there are currently no bioherbicides commercially available against barnyardgrass. Several species of Bipolaris have been studied extensively for weed control. Bipolaris sorghicola and an unidentified Bipolaris species showed efficacy against Johnson grass (Sorghum halepense; Winder & Van Dyke, 1990), while an isolate of B. setariae was highly pathogenic on goosegrass (Eleusine indica; Figliola, Camper, *Corresponding author. Email: [email protected] © 2014 Taylor & Francis 840 J. Zhang et al. & Ridings, 1988). An isolate of B. sacchari was found to cause a leaf-spot disease on climbing ferns (Lygodium microphyllum) and was used successfully for control of cogongrass (Imperata cylindrica; Yandoc, Charudattan, & Shilling, 2005). However, no previous investigations of the effects of Bipolaris eleusines Alcorn & R.G. Shivas on weeds have been reported. The objectives of this study were to: (1) verify the identification of the most efficacious B. eleusines isolate using rDNA sequencing; (2) assess the potential of this B. eleusines isolate as a mycoherbicide candidate by studying the effect on weed growth stage, fungal inoculum dose and an emulsion formulation on weed-control efficacy; and (3) understand the risk and value of this isolate based on its impact on common crop species in China and weeds in paddy rice fields. An isolate of B. eleusines, obtained originally from severely diseased barnyard- grass plants in rice fields of Guangxi province, China by Yu et al. (2005), was used. Its pathogenicity on barnyardgrass was confirmed by the same authors based on the Koch’s postulates. The isolate has been stored on potato dextrose agar (PDA) at 4°C at the Weed Laboratory of the China National Rice Research Institute (CNRRI), with periodic transfer, inoculation of barnyardgrass and reisolation from leaf lesions on the host. The original identification of the fungus was based on the disease symptoms and fungal morphology. To verify the taxonomic designa- tion, rDNA sequence of the isolate was used to compare with that of B. eleusines at GenBank (http://blast.ncbi.nlm.nih.gov). The fungus was cultured in potato dex‐ trose broth at 28°C for two days in darkness. Mycelia were collected by filtering the fungal culture through two layers of cheesecloth, washing twice with sterile distilled water and drying on filter paper. The total genomic DNA of B. eleusines was extracted using a cell-SDS-lysis protocol described originally by Graham, Mayers, and Henry (1994). The polymerase chain reaction (PCR) products of 18S rDNA, ITS and 5.8S rDNA were obtained with the universal primers NS1 and ITS2 (White, Bruns, Lee, & Taylor, 1990). Sequencing was done using the ABI Big-dye Ready-Reaction Kit (Applied Biosystems Inc., USA) with a one-eighth reaction, and sequences were generated on an ABI 3730 XL Sequencer (Applied Biosystems Inc., USA). ITS sequences of other B. eleusines isolates (GenBank accession numbers: AB179837, AF071326, AF081452, AF163070, AF071318, AF081451, AF163071, AF158109, Downloaded by [Jianping Zhang] at 18:36 19 May 2014 AY004788, AY004801, AY004796, DQ300207, DQ337380, GQ870276 and GU256748) were obtained from GenBank for comparisons with the current isolate. Multiple alignments were performed using ClustalX1.83 (Thompson, Gibson, Plewniak, Jeanmougin, & Higgins, 1997) and refined manually. Adjustments of sequence alignments were performed using the data editor program Mega 3.1 software (Kumar, Tamura, & Nei, 2004). Following alignment, phylogenetic trees were constructed by Neighbor-joining (NJ) with Mega 3.1. Statistical support was evaluated by bootstrap analysis with 1000 replications. To prepare the fungal inoculum, a 7-mm mycelial disc from the margin of an actively growing colony of the isolate on PDA was transferred to PDA and grown for 10–14 days in darkness at 28°C until sporulation. The conidia were harvested by flooding plates with 0.05% Tween-20 (V/V) and scraping. After filtering through two layers of cheesecloth, conidial suspensions were adjusted to various concentrations with sterile distilled water using a haemocytometer. Biocontrol Science and Technology 841 Seeds of barnyardgrass were collected from rice fields at the Fuyang Experiment Farm of CNRRI, Hangzhou, China, and scarified by soaking in 0.1% nitric acid (KNO3) for 30 min prior to planting. Rice seeds [indica rice (Jiayu293), japonica rice (Xiushui11) and
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