Niclosamide inhibits leaf blight caused by Xanthomonas oryzae in rice
Identifieur interne : 000060 ( Pmc/Corpus ); précédent : 000059; suivant : 000061Niclosamide inhibits leaf blight caused by Xanthomonas oryzae in rice
Auteurs : Sung-Il Kim ; Jong Tae Song ; Jin-Yong Jeong ; Hak Soo SeoSource :
- Scientific Reports [ 2045-2322 ] ; 2016.
Abstract
Rice leaf blight, which is caused by the bacterial pathogen
Url:
DOI: 10.1038/srep21209
PubMed: 26879887
PubMed Central: 4754756
Links to Exploration step
PMC:4754756Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Niclosamide inhibits leaf blight caused by <italic>Xanthomonas oryzae</italic> in rice</title><author><name sortKey="Kim, Sung Il" sort="Kim, Sung Il" uniqKey="Kim S" first="Sung-Il" last="Kim">Sung-Il Kim</name><affiliation><nlm:aff id="a1"><institution>Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation></author><author><name sortKey="Song, Jong Tae" sort="Song, Jong Tae" uniqKey="Song J" first="Jong Tae" last="Song">Jong Tae Song</name><affiliation><nlm:aff id="a2"><institution>School of Applied Biosciences, Kyungpook National University</institution>, Daegu 702-701,<country>Korea</country></nlm:aff></affiliation></author><author><name sortKey="Jeong, Jin Yong" sort="Jeong, Jin Yong" uniqKey="Jeong J" first="Jin-Yong" last="Jeong">Jin-Yong Jeong</name><affiliation><nlm:aff id="a3"><institution>Department of Convergence Medicine and Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine</institution>, Seoul 05505,<country>Republic of Korea</country></nlm:aff></affiliation></author><author><name sortKey="Seo, Hak Soo" sort="Seo, Hak Soo" uniqKey="Seo H" first="Hak Soo" last="Seo">Hak Soo Seo</name><affiliation><nlm:aff id="a1"><institution>Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation><affiliation><nlm:aff id="a4"><institution>Plant Genomics and Breeding Institute, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation><affiliation><nlm:aff id="a5"><institution>Bio-MAX Institute, Seoul National University</institution>, Seoul 151-818,<country>Korea</country></nlm:aff></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">26879887</idno><idno type="pmc">4754756</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4754756</idno><idno type="RBID">PMC:4754756</idno><idno type="doi">10.1038/srep21209</idno><date when="2016">2016</date><idno type="wicri:Area/Pmc/Corpus">000060</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000060</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Niclosamide inhibits leaf blight caused by <italic>Xanthomonas oryzae</italic> in rice</title><author><name sortKey="Kim, Sung Il" sort="Kim, Sung Il" uniqKey="Kim S" first="Sung-Il" last="Kim">Sung-Il Kim</name><affiliation><nlm:aff id="a1"><institution>Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation></author><author><name sortKey="Song, Jong Tae" sort="Song, Jong Tae" uniqKey="Song J" first="Jong Tae" last="Song">Jong Tae Song</name><affiliation><nlm:aff id="a2"><institution>School of Applied Biosciences, Kyungpook National University</institution>, Daegu 702-701,<country>Korea</country></nlm:aff></affiliation></author><author><name sortKey="Jeong, Jin Yong" sort="Jeong, Jin Yong" uniqKey="Jeong J" first="Jin-Yong" last="Jeong">Jin-Yong Jeong</name><affiliation><nlm:aff id="a3"><institution>Department of Convergence Medicine and Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine</institution>, Seoul 05505,<country>Republic of Korea</country></nlm:aff></affiliation></author><author><name sortKey="Seo, Hak Soo" sort="Seo, Hak Soo" uniqKey="Seo H" first="Hak Soo" last="Seo">Hak Soo Seo</name><affiliation><nlm:aff id="a1"><institution>Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation><affiliation><nlm:aff id="a4"><institution>Plant Genomics and Breeding Institute, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></nlm:aff></affiliation><affiliation><nlm:aff id="a5"><institution>Bio-MAX Institute, Seoul National University</institution>, Seoul 151-818,<country>Korea</country></nlm:aff></affiliation></author></analytic><series><title level="j">Scientific Reports</title><idno type="eISSN">2045-2322</idno><imprint><date when="2016">2016</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Rice leaf blight, which is caused by the bacterial pathogen <italic>Xanthomonas oryzae</italic> pv<italic>. oryzae</italic> (<italic>Xoo</italic>), results in huge losses in grain yield. Here, we show that <italic>Xoo</italic>-induced rice leaf blight is effectively controlled by niclosamide, an oral antihelminthic drug and molluscicide, which also functions as an anti-tumor agent. Niclosamide directly inhibited the growth of the three <italic>Xoo</italic> strains PXO99, 10208 and K3a. Niclosamide moved long distances from the site of local application to distant rice tissues. Niclosamide also increased the levels of salicylate and induced the expression of defense-related genes such as <italic>OsPR1</italic> and <italic>OsWRKY45</italic>, which suppressed <italic>Xoo</italic>-induced leaf wilting. Niclosamide had no detrimental effects on vegetative/reproductive growth and yield. These combined results indicate that niclosamide can be used to block bacterial leaf blight in rice with no negative side effects.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Mew, T W" uniqKey="Mew T">T. W. Mew</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Tabei, H" uniqKey="Tabei H">H. Tabei</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ishiyama, S" uniqKey="Ishiyama S">S. Ishiyama</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ray, S K" uniqKey="Ray S">S. K. Ray</name></author><author><name sortKey="Rajeshwari, R" uniqKey="Rajeshwari R">R. Rajeshwari</name></author><author><name sortKey="Sonti, R V" uniqKey="Sonti R">R. V. Sonti</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Koplin, R" uniqKey="Koplin R">R. Köplin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hu, J" uniqKey="Hu J">J. Hu</name></author><author><name sortKey="Qian, W" uniqKey="Qian W">W. Qian</name></author><author><name sortKey="He, C" uniqKey="He C">C. He</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Rajeshwari, R" uniqKey="Rajeshwari R">R. Rajeshwari</name></author><author><name sortKey="Jha, G" uniqKey="Jha G">G. Jha</name></author><author><name sortKey="Sonti, R V" uniqKey="Sonti R">R. V. Sonti</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jha, G" uniqKey="Jha G">G. Jha</name></author><author><name sortKey="Rajeshwari, R" uniqKey="Rajeshwari R">R. Rajeshwari</name></author><author><name sortKey="Sonti, R V" uniqKey="Sonti R">R. V. Sonti</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chithrashree, A C" uniqKey="Chithrashree A">A. C. Chithrashree</name></author><author><name sortKey="Udayashankar, S" uniqKey="Udayashankar S">S. Udayashankar</name></author><author><name sortKey="Chandra Nayaka, M S" uniqKey="Chandra Nayaka M">M. S. Chandra Nayaka</name></author><author><name sortKey="Srinivas, R C" uniqKey="Srinivas R">R. C. Srinivas</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gnanamanickam, S S" uniqKey="Gnanamanickam S">S. S. Gnanamanickam</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chand, T" uniqKey="Chand T">T. Chand</name></author><author><name sortKey="Sing, N" uniqKey="Sing N">N. Sing</name></author><author><name sortKey="Sing, H" uniqKey="Sing H">H. Sing</name></author><author><name sortKey="Thind, B S" uniqKey="Thind B">B. S. Thind</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Khan, J A" uniqKey="Khan J">J. A. Khan</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Karthikeyan, V" uniqKey="Karthikeyan V">V. Karthikeyan</name></author><author><name sortKey="Gnanamanickam, S S" uniqKey="Gnanamanickam S">S. S. Gnanamanickam</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Iwai, T" uniqKey="Iwai T">T. Iwai</name></author><author><name sortKey="Seo, S" uniqKey="Seo S">S. Seo</name></author><author><name sortKey="Mitsuhara, I" uniqKey="Mitsuhara I">I. Mitsuhara</name></author><author><name sortKey="Ohashi, Y" uniqKey="Ohashi Y">Y. Ohashi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Mcmanus, P S" uniqKey="Mcmanus P">P. S. McManus</name></author><author><name sortKey="Stockwell, V O" uniqKey="Stockwell V">V. O. Stockwell</name></author><author><name sortKey="Sundin, G W" uniqKey="Sundin G">G. W. Sundin</name></author><author><name sortKey="Jones, A L" uniqKey="Jones A">A. L. Jones</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Bhasin, H" uniqKey="Bhasin H">H. Bhasin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Natrajkumar, P" uniqKey="Natrajkumar P">P. Natrajkumar</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Song, W Y" uniqKey="Song W">W. Y. Song</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Yang, D" uniqKey="Yang D">D. Yang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gu, K Y" uniqKey="Gu K">K. Y. Gu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Liu, D O" uniqKey="Liu D">D. O. Liu</name></author><author><name sortKey="Ronald, P C" uniqKey="Ronald P">P. C. Ronald</name></author><author><name sortKey="Bogdanove, A J" uniqKey="Bogdanove A">A. J. Bogdanove</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Cheema, K K" uniqKey="Cheema K">K. K. Cheema</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Yoshimura, S" uniqKey="Yoshimura S">S. Yoshimura</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hulbert, S H" uniqKey="Hulbert S">S. H. Hulbert</name></author><author><name sortKey="Webb, C A" uniqKey="Webb C">C. A. Webb</name></author><author><name sortKey="Smith, S M" uniqKey="Smith S">S. M. Smith</name></author><author><name sortKey="Sun, Q" uniqKey="Sun Q">Q. Sun</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="White, F F" uniqKey="White F">F. F. White</name></author><author><name sortKey="Yang, B" uniqKey="Yang B">B. Yang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Verdier, V" uniqKey="Verdier V">V. Verdier</name></author><author><name sortKey="Cruz, C V" uniqKey="Cruz C">C. V. Cruz</name></author><author><name sortKey="Leach, J E" uniqKey="Leach J">J. E. Leach</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Singh, S" uniqKey="Singh S">S. Singh</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lia, Q" uniqKey="Lia Q">Q. Lia</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Cernadas, R A" uniqKey="Cernadas R">R. A. Cernadas</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Sharma, A" uniqKey="Sharma A">A. Sharma</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Swamy, P" uniqKey="Swamy P">P. Swamy</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Han, M" uniqKey="Han M">M. Han</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Peng, X" uniqKey="Peng X">X. Peng</name></author></analytic></biblStruct><biblStruct></biblStruct><biblStruct><analytic><author><name sortKey="Nakayama, A" uniqKey="Nakayama A">A. Nakayama</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Goto, S" uniqKey="Goto S">S. Goto</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wu, C J" uniqKey="Wu C">C. J. Wu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zhu, P J" uniqKey="Zhu P">P. J. Zhu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Osada, T" uniqKey="Osada T">T. Osada</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Balgi, A D" uniqKey="Balgi A">A. D. Balgi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chen, M" uniqKey="Chen M">M. Chen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wang, A M" uniqKey="Wang A">A. M. Wang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Fonseca, B D" uniqKey="Fonseca B">B. D. Fonseca</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Weinbach, E C" uniqKey="Weinbach E">E. C. Weinbach</name></author><author><name sortKey="Garbus, J" uniqKey="Garbus J">J. Garbus</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Prathuangwong, S" uniqKey="Prathuangwong S">S. Prathuangwong</name></author><author><name sortKey="Amnuaykit, K" uniqKey="Amnuaykit K">K. Amnuaykit</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Mitsuhara, I" uniqKey="Mitsuhara I">I. Mitsuhara</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hwang, S H" uniqKey="Hwang S">S.-H. Hwang</name></author><author><name sortKey="Lee, I A" uniqKey="Lee I">I. A. Lee</name></author><author><name sortKey="Yie, S W" uniqKey="Yie S">S. W. Yie</name></author><author><name sortKey="Hwang, D J" uniqKey="Hwang D">D.-J. Hwang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ryu, H S" uniqKey="Ryu H">H.-S. Ryu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Liua, X" uniqKey="Liua X">X. Liua</name></author><author><name sortKey="Baia, X" uniqKey="Baia X">X. Baia</name></author><author><name sortKey="Wanga, X" uniqKey="Wanga X">X. Wanga</name></author><author><name sortKey="Chua, C" uniqKey="Chua C">C. Chua</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jin, Y" uniqKey="Jin Y">Y. Jin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Sack, U" uniqKey="Sack U">U. Sack</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Vleesschauwer, D D" uniqKey="Vleesschauwer D">D. D. Vleesschauwer</name></author><author><name sortKey="Xu, J" uniqKey="Xu J">J. Xu</name></author><author><name sortKey="Hofte, M" uniqKey="Hofte M">M. Höfte</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Tao, Z" uniqKey="Tao Z">Z. Tao</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Iwata, M" uniqKey="Iwata M">M. Iwata</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Friedrich, L" uniqKey="Friedrich L">L. Friedrich</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gorlach, J" uniqKey="Gorlach J">J. Görlach</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lawton, K A" uniqKey="Lawton K">K. A. Lawton</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Francesco, I" uniqKey="Francesco I">I. Francesco</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ofori Adjei, D" uniqKey="Ofori Adjei D">D. Ofori-Adjei</name></author><author><name sortKey="Dodoo, A N O" uniqKey="Dodoo A">A. N. O. Dodoo</name></author><author><name sortKey="Appiah Danquah, A" uniqKey="Appiah Danquah A">A. Appiah-Danquah</name></author><author><name sortKey="Couper, M" uniqKey="Couper M">M. Couper</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Park, B S" uniqKey="Park B">B. S. Park</name></author><author><name sortKey="Song, J T" uniqKey="Song J">J. T. Song</name></author><author><name sortKey="Seo, H S" uniqKey="Seo H">H. S. Seo</name></author></analytic></biblStruct></listBibl></div1></back></TEI><pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">Sci Rep</journal-id><journal-id journal-id-type="iso-abbrev">Sci Rep</journal-id><journal-title-group><journal-title>Scientific Reports</journal-title></journal-title-group><issn pub-type="epub">2045-2322</issn><publisher><publisher-name>Nature Publishing Group</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">26879887</article-id><article-id pub-id-type="pmc">4754756</article-id><article-id pub-id-type="pii">srep21209</article-id><article-id pub-id-type="doi">10.1038/srep21209</article-id><article-categories><subj-group subj-group-type="heading"><subject>Article</subject></subj-group></article-categories><title-group><article-title>Niclosamide inhibits leaf blight caused by <italic>Xanthomonas oryzae</italic> in rice</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Kim</surname><given-names>Sung-Il</given-names></name><xref ref-type="aff" rid="a1">1</xref></contrib><contrib contrib-type="author"><name><surname>Song</surname><given-names>Jong Tae</given-names></name><xref ref-type="aff" rid="a2">2</xref></contrib><contrib contrib-type="author"><name><surname>Jeong</surname><given-names>Jin-Yong</given-names></name><xref ref-type="aff" rid="a3">3</xref></contrib><contrib contrib-type="author"><name><surname>Seo</surname><given-names>Hak Soo</given-names></name><xref ref-type="corresp" rid="c1">a</xref><xref ref-type="aff" rid="a1">1</xref><xref ref-type="aff" rid="a4">4</xref><xref ref-type="aff" rid="a5">5</xref></contrib><aff id="a1"><label>1</label><institution>Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></aff><aff id="a2"><label>2</label><institution>School of Applied Biosciences, Kyungpook National University</institution>, Daegu 702-701,<country>Korea</country></aff><aff id="a3"><label>3</label><institution>Department of Convergence Medicine and Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine</institution>, Seoul 05505,<country>Republic of Korea</country></aff><aff id="a4"><label>4</label><institution>Plant Genomics and Breeding Institute, Seoul National University</institution>, Seoul 151-921,<country>Korea</country></aff><aff id="a5"><label>5</label><institution>Bio-MAX Institute, Seoul National University</institution>, Seoul 151-818,<country>Korea</country></aff></contrib-group><author-notes><corresp id="c1"><label>a</label><email>seohs@snu.ac.kr</email></corresp></author-notes><pub-date pub-type="epub"><day>16</day><month>02</month><year>2016</year></pub-date><pub-date pub-type="collection"><year>2016</year></pub-date><volume>6</volume><elocation-id>21209</elocation-id><history><date date-type="received"><day>18</day><month>09</month><year>2015</year></date><date date-type="accepted"><day>19</day><month>01</month><year>2016</year></date></history><permissions><copyright-statement>Copyright © 2016, Macmillan Publishers Limited</copyright-statement><copyright-year>2016</copyright-year><copyright-holder>Macmillan Publishers Limited</copyright-holder><license license-type="open-access" xlink:href="http://creativecommons.org/licenses/by/4.0/"><pmc-comment>author-paid</pmc-comment>
<license-p>This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit <ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">http://creativecommons.org/licenses/by/4.0/</ext-link></license-p></license></permissions><abstract><p>Rice leaf blight, which is caused by the bacterial pathogen <italic>Xanthomonas oryzae</italic> pv<italic>. oryzae</italic> (<italic>Xoo</italic>), results in huge losses in grain yield. Here, we show that <italic>Xoo</italic>-induced rice leaf blight is effectively controlled by niclosamide, an oral antihelminthic drug and molluscicide, which also functions as an anti-tumor agent. Niclosamide directly inhibited the growth of the three <italic>Xoo</italic> strains PXO99, 10208 and K3a. Niclosamide moved long distances from the site of local application to distant rice tissues. Niclosamide also increased the levels of salicylate and induced the expression of defense-related genes such as <italic>OsPR1</italic> and <italic>OsWRKY45</italic>, which suppressed <italic>Xoo</italic>-induced leaf wilting. Niclosamide had no detrimental effects on vegetative/reproductive growth and yield. These combined results indicate that niclosamide can be used to block bacterial leaf blight in rice with no negative side effects.</p></abstract></article-meta></front><body><p>Bacterial blight is one of the most serious diseases of rice and is especially prevalent in irrigated and rain-fed lowland areas. Bacterial blight has the following three symptoms: leaf blight, kresek (seedling blight or wilt phase of the syndrome), and pale-yellow leaves<xref ref-type="bibr" rid="b1">1</xref>. This disease is sometimes referred to as bacterial leaf blight, indicating that the leaf blight phase of the syndrome is the most distinct and commonly observed phase<xref ref-type="bibr" rid="b1">1</xref>. The most common symptom of this disease is leaf wilting, especially in young leaves, because the bacterial blight pathogen multiplies in xylem elements and primarily invades the vascular tissue<xref ref-type="bibr" rid="b2">2</xref>.</p><p>Bacterial blight in rice is caused by <italic>Xanthomonas oryzae</italic> pv<italic>. oryzae</italic> (<italic>Xoo</italic>)<xref ref-type="bibr" rid="b3">3</xref> and results in huge losses in grain yield. <italic>Xoo</italic> is a phytopathogenic Gram-negative bacterium that belongs to the family <italic>Pseudomonadaceae. Xoo</italic> enters the plant through wounds or hydathodes, multiplies in the epitheme, moves to the xylem vessels, and undergoes active multiplication, which results in blight disease symptoms in rice leaves. <italic>Xoo</italic> produces a range of virulence factors, including exopolysaccharides, extracellular enzymes, iron-chelating siderophores, and type III secretion-dependent effectors, which are collectively essential for virulence<xref ref-type="bibr" rid="b4">4</xref><xref ref-type="bibr" rid="b5">5</xref><xref ref-type="bibr" rid="b6">6</xref><xref ref-type="bibr" rid="b7">7</xref><xref ref-type="bibr" rid="b8">8</xref>.</p><p>Numerous studies have investigated rice protection from bacterial blight. Three different strategies, biological control, chemical control, and genetic resistance, have been employed to manage plant diseases such as bacterial leaf blight. Specific bacterial strains have been used for biological control of rice leaf blight; for example, fresh suspensions and powdered formulations of plant growth-promoting rhizobacteria<xref ref-type="bibr" rid="b9">9</xref>. <italic>Bacillus</italic> spp. and <italic>Pseudomonas</italic> spp. also have been used to suppress rice leaf blight<xref ref-type="bibr" rid="b10">10</xref>. However, pathogen variation and the lack of suitable biological agents limit the efficacy of biological controls, although such measures remain a suitable method for disease control and management because they are environmentally friendly.</p><p>Numerous chemicals have been tested for chemical control of bacterial leaf blight. For example, leaf blight lesions in rice were reduced by application of bleaching powder containing 30% chlorine<xref ref-type="bibr" rid="b11">11</xref>. The broad-spectrum antibiotics benzylpenicillin, ampicillin, kanamycin, streptomycin, chloramphenicol, and sinobionic also inhibit the growth of <italic>Xoo</italic> isolates, although their effects were examined only <italic>in vitro</italic><xref ref-type="bibr" rid="b12">12</xref>. Benzothiadiazole, a known plant activator, exerts protective effects against a broad spectrum of diseases including bacterial leaf blight, without inducing major adverse effects on plant growth when applied at appropriate dosages<xref ref-type="bibr" rid="b13">13</xref>. Probenazole also protects rice from blast caused by the fungal pathogen <italic>Magnaporthe grisea</italic><xref ref-type="bibr" rid="b14">14</xref>. Streptomycin has been used for crop protection from bacterial and fungal diseases, including fire blight, bacterial blight, and soft rot<xref ref-type="bibr" rid="b15">15</xref>. Oxytetracycline, gentamycin, and oxolinic acid have been used to control several bacterial and fungal diseases<xref ref-type="bibr" rid="b15">15</xref>. However, an effective and economical chemical treatment for rice leaf blight has not been established, although research and development are ongoing.</p><p>Enhancing plant genetic resistance is an effective method for controlling bacterial leaf blight disease. A number of studies have identified plant genes that confer resistance against <italic>Xanthomonas</italic> bacteria. At least 38 bacterial leaf blight resistance gene (<italic>R</italic> genes), designated in series from <italic>Xa1</italic> to <italic>Xa38</italic>, have been identified<xref ref-type="bibr" rid="b16">16</xref><xref ref-type="bibr" rid="b17">17</xref>. Six of these genes have been cloned (<italic>Xa1</italic>, <italic>xa5</italic>, <italic>xa13</italic>, <italic>Xa21</italic>, <italic>Xa3</italic>/<italic>Xa26</italic>, and <italic>Xa27</italic>), and six additional genes have been physically mapped (<italic>Xa2</italic>, <italic>Xa4</italic>, <italic>Xa7</italic>, <italic>Xa30</italic>, <italic>Xa33</italic>, and <italic>Xa38</italic>)<xref ref-type="bibr" rid="b16">16</xref><xref ref-type="bibr" rid="b17">17</xref><xref ref-type="bibr" rid="b18">18</xref><xref ref-type="bibr" rid="b19">19</xref><xref ref-type="bibr" rid="b20">20</xref><xref ref-type="bibr" rid="b21">21</xref><xref ref-type="bibr" rid="b22">22</xref>. Two major classes of <italic>R</italic> genes, receptor kinase (RLK) and nucleotide-binding site leucine rice repeat (NBS)-LRR, are involved in disease resistance in rice. <italic>Xa21</italic> is the first <italic>R</italic> gene of the RLK class to be cloned with a broad spectrum of resistance, and <italic>Xa1</italic> is an <italic>R</italic> gene of the NBS-LRR class<xref ref-type="bibr" rid="b23">23</xref>, which is the largest <italic>R</italic> gene class conferring resistance against bacteria, fungi, and viruses<xref ref-type="bibr" rid="b24">24</xref>. Other <italic>R</italic> genes, including <italic>xa5</italic> and <italic>xa13</italic>, encode proteins such as a small subunit of the transcription factor IIA (TFIIAγ) and a plasma membrane protein, respectively<xref ref-type="bibr" rid="b25">25</xref><xref ref-type="bibr" rid="b26">26</xref>. These data have been used to develop a breeding program for bacterial leaf blight-resistant rice. The introduction of resistance genes <italic>xa5</italic>, <italic>xa13</italic>, and <italic>Xa21</italic> into new cultivars conferred a high level of resistance against <italic>Xoo</italic><xref ref-type="bibr" rid="b27">27</xref>, indicating that germplasm screening against bacterial leaf blight and breeding-mediated transfer of resistant genes to target cultivars can generate new rice cultivars with increased resistance to bacterial leaf blight disease. Large-scale experiments, including microarrays, were used to isolate genes related to bacterial leaf blight; a large number of candidate genes was selected, although their functions were not clearly identified<xref ref-type="bibr" rid="b28">28</xref><xref ref-type="bibr" rid="b29">29</xref>. A transgenic approach was employed to protect rice from bacterial leaf blight and fungal blast. For example, transgenic rice overexpressing cecropin B (an antibacterial peptide from <italic>Bombyx mori</italic>) showed increased resistance to bacterial leaf blight<xref ref-type="bibr" rid="b30">30</xref>. The introduction of multiple bacterial blight resistance genes (<italic>Xa4</italic>, <italic>xa5</italic>, <italic>xa13</italic>, and <italic>Xa21</italic>) into rice conferred resistance to six <italic>Xoo</italic> bacteria<xref ref-type="bibr" rid="b31">31</xref>. Rice transformants overexpressing WRKY30 are resistant to <italic>Xoo</italic><xref ref-type="bibr" rid="b32">32</xref> and the fungal pathogen <italic>Magnaporthe oryzae</italic><xref ref-type="bibr" rid="b33">33</xref>, and those overexpressing WRKY45 displayed markedly enhanced resistance to bacterial leaf blight and fungal blast disease<xref ref-type="bibr" rid="b34">34</xref><xref ref-type="bibr" rid="b35">35</xref><xref ref-type="bibr" rid="b36">36</xref>. These results suggest that transgenic approaches can be effectively utilized to develop disease-resistant rice.</p><p>Although several biological, chemical, and genetic control approaches have yielded improvements in rice protection from bacterial leaf blight, there was still a need for an effective approach that would provide large-scale protection. Therefore, we focused our current efforts on screening known chemicals for their effects on rice leaf blight. We are interested in the discovery of a master regulator that effectively functions to protect animals and plants from infectious diseases mediated by bacteria, fungi, and/or parasites. Therefore, we first considered infectious diseases of humans. We selected two drugs that cure infectious diseases in humans, auranofin [3,4,5-triacetyloxy-6-(acetyloxymethyl)oxane-2-thiolate] and niclosamide [5-chloro-<italic>N</italic>-(2-chloro-4-nitrophenyl)-2-hydroxybenzamide], for further analysis. However, because auranofin is much more expensive than niclosamide, we focused our initial screen on niclosamide. Niclosamide has been widely used since 1960 to treat gastrointestinal tapeworm infections in both humans and animals. Recent studies show that niclosamide has antiviral activity against the severe acute respiratory syndrome virus<xref ref-type="bibr" rid="b37">37</xref>, anti-anthrax toxin properties<xref ref-type="bibr" rid="b38">38</xref>, and anti-neoplastic activity<xref ref-type="bibr" rid="b39">39</xref>. Niclosamide strongly induces LC3-positive autophagosomes<xref ref-type="bibr" rid="b40">40</xref>, inhibits the Wnt/Frizzled pathway<xref ref-type="bibr" rid="b41">41</xref>, suppresses the autonomous notch-signaling pathway<xref ref-type="bibr" rid="b42">42</xref>, and inhibits mTOR signaling<xref ref-type="bibr" rid="b43">43</xref>. Niclosamide uncouples mitochondrial oxidative phosphorylation<xref ref-type="bibr" rid="b44">44</xref> and thereby slows cell growth. These combined results indicate that niclosamide has various curative effects on humans and animals. We reasoned that niclosamide might affect plant disease responses by modulating disease signaling pathways as a broad spectrum regulator. Therefore, we investigated the effect of niclosamide on bacterial leaf blight in rice.</p><p>Here, we show that niclosamide blocks rice leaf wilting mediated by <italic>Xoo</italic> bacteria, both locally and systemically, without negatively affecting plant growth. The results suggest that niclosamide may be used to prevent rice leaf blight caused by bacterial pathogen attack.</p><sec disp-level="1"><title>Results</title><sec disp-level="2"><title>Niclosamide inhibits <italic>Xoo</italic> bacterial growth</title><p>To evaluate the functional effect of niclosamide on bacterial blight, we first examined its effect on the growth of <italic>Xoo</italic> bacteria using three different strains, PXO99, 10208, and K3a, and <italic>Xanthomonas axonopodis</italic> pv. <italic>glycines</italic> (<italic>Xag</italic>). The results showed that growth of the <italic>Xoo</italic> strains was completely inhibited by 5 μg/ml niclosamide, whereas <italic>Xag</italic> growth was inhibited by 15 μg/ml niclosamide (<xref ref-type="fig" rid="f1">Fig. 1a</xref>). <italic>Xag</italic> is a pathogen that causes bacterial leaf pustule disease in soybean<xref ref-type="bibr" rid="b45">45</xref>. We also examined the effect of niclosamide on the growth of the fungal pathogen <italic>Magnaporthe oryzae</italic>. Niclosamide did exert an inhibitory effect on the growth of 12 <italic>M. oryzae</italic> strains, although the inhibition was considerably weaker than that against <italic>Xoo</italic> (<xref ref-type="fig" rid="f1">Fig. 1b</xref>).</p><p>We also tested the effects of parthenolide, a sesquiterpene lactone with anti-tumor activity, on the growth of the <italic>Xoo</italic> strains PXO99 and 10208. Parthenolide failed to inhibit the growth of either <italic>Xoo</italic> strain at a concentration of 5 μg/ml, although it slightly inhibited bacterial growth at 50 μg/ml (<xref ref-type="fig" rid="f1">Fig. 1c</xref>). We also tested the effect of niclosamide on the growth of three <italic>E. coli</italic> strains, Top10, Rosseta2, and DH10b, which served as Gram-negative control bacteria. At lower concentrations, niclosamide had no effect on the growth of these <italic>E. coli</italic> strains, although it slightly inhibited growth at 50 μg/ml (<xref ref-type="fig" rid="f1">Fig. 1d</xref>).</p><p>To determine the minimum inhibitory concentration (MIC) of niclosamide on PXO99 and 10208 growth, we tested concentrations ranging from 0–5 μg/ml. Both <italic>Xoo</italic> strains could grow in the presence of 4 μg/ml niclosamide, but not 5 μg/ml niclosamide (<xref ref-type="fig" rid="f1">Fig. 1e</xref>; left panel). We further narrowed the MIC of niclosamide from 4–5 μg/ml. As shown in <xref ref-type="fig" rid="f1">Fig. 1e</xref>, the growth of both PXO99 and 10208 was completely inhibited by 4.2 μg/ml niclosamide (<xref ref-type="fig" rid="f1">Fig. 1e</xref>; right panel), indicating that 4.2 μg/ml niclosamide is the MIC for both <italic>Xoo</italic> strains.</p></sec><sec disp-level="2"><title>Rice bacterial blight is blocked by niclosamide</title><p>We examined whether niclosamide blocks bacterial blight in rice. For this experiment, we grew the rice cultivar Nipponbare in a growth chamber for 12 weeks (before bolting), and subjected the plants to pathogen and niclosamide treatment.</p><p>First, to determine the minimum concentration of niclosamide that would block bacterial blight, we examined the niclosamide dosage effect on disease responses to the representative <italic>Xoo</italic> strain PXO99. We inoculated PXO99 onto Nipponbare using the leaf-clipping method, sprayed the plants with different niclosamide concentrations, and examined the phenotypes of plants treated with pathogen only or pathogen plus niclosamide. Leaf wilting did not develop in plants treated with ≥8 μg/ml niclosamide (<xref ref-type="fig" rid="f2">Fig. 2a</xref>). We also estimated lesion development by measuring lesion length. Lesion development also was significantly inhibited by 8 μg/ml niclosamide, although a small lesion was still detected at this concentration (<xref ref-type="fig" rid="f2">Fig. 2b</xref>). Lesion development gradually declined with increasing niclosamide concentrations (<xref ref-type="fig" rid="f2">Fig. 2b</xref>). We also examined the levels of bacterial growth, and found that population levels of <italic>Xoo</italic> bacteria declined with increasing niclosamide concentrations (<xref ref-type="fig" rid="f2">Fig. 2c</xref>).</p><p>We next examined the effect of niclosamide on rice disease responses to <italic>Xoo</italic> bacteria after treatment with PXO99. First, Nipponbare rice was inoculated with PXO99 by the leaf-clipping method. After incubation for four days until leaf blight lesions with a length of 2 cm developed, the plants were sprayed with 8 μg/ml niclosamide (<xref ref-type="fig" rid="f3">Fig. 3</xref>). Leaf blight was completely blocked in niclosamide-treated leaves (<xref ref-type="fig" rid="f3">Fig. 3a</xref>). Lesion development also was completely blocked in niclosamide-treated leaves (<xref ref-type="fig" rid="f3">Fig. 3b</xref>), and PXO99 growth was inhibited (<xref ref-type="fig" rid="f3">Fig. 3c</xref>).</p></sec><sec disp-level="2"><title>Niclosamide has a systemic effect on rice disease response</title><p>We next investigated whether niclosamide could move from the site of local application to distant tissues and subsequently inhibit PXO99-mediated leaf wilting. We inoculated half of the leaves of Nipponbare plants with PXO99 and covered them with polythene bags. Then, we sprayed the non-inoculated leaves with niclosamide and examined leaf blight in both local and systemic leaves. Leaf wilting and lesion development were completely inhibited in PXO99-inoculated leaves that had not been treated with niclosamide (<xref ref-type="fig" rid="f4">Fig. 4a,b</xref>). PXO99 growth also was significantly inhibited in the inoculated leaves (<xref ref-type="fig" rid="f4">Fig. 4c</xref>).</p><p>Next, we examined the long-distance movement of niclosamide by extracting niclosamide from niclosamide-treated leaves and untreated systemic leaves. Niclosamide levels gradually increased in systemic leaves (<xref ref-type="fig" rid="f5">Fig. 5a,b</xref>), indicating that niclosamide can systemically move from niclosamide-treated local leaves to untreated distal leaves.</p></sec><sec disp-level="2"><title>Niclosamide induces the expression of defense-related genes</title><p>We next examined whether niclosamide protects PXO99-infected plants through the induction of pathogen-related gene expression. We treated the rice cultivar Nipponbare with different niclosamide concentrations for 24 h, and then extracted total RNA from niclosamide-treated and untreated control plants. We performed qRT-PCR analysis to examine the transcript levels of seven defense-related genes, including <italic>OsPR1</italic>, <italic>OsPR10</italic>, <italic>OsWRKY45</italic>, <italic>OsWRKY62</italic>, <italic>OsWRKY71</italic>, <italic>OsHI-LOX</italic>, and <italic>OsACS2</italic>. The results showed that <italic>OsPR1</italic>, <italic>OsPR10</italic>, <italic>OsWRKY45</italic>, <italic>OsWRKY62</italic>, <italic>OsWRKY71</italic>, and <italic>OsHI-LOX</italic> were induced by niclosamide treatment, and that transcript expression levels increased with increasing niclosamide levels (<xref ref-type="fig" rid="f6">Fig. 6</xref>). However, <italic>OsACS2</italic> expression was only slightly increased 30 min after treatment with 40 μg/ml niclosamide, which suggests that its expression was not affected by niclosamide (<xref ref-type="fig" rid="f6">Fig. 6</xref>).</p><p>The pathogenesis-related genes <italic>OsPR1</italic>, <italic>OsPR10</italic>, <italic>OsWRKY45</italic>, <italic>OsWRKY62</italic>, and <italic>OsWRKY71</italic> are induced by high salicylate (SA) levels<xref ref-type="bibr" rid="b46">46</xref><xref ref-type="bibr" rid="b47">47</xref><xref ref-type="bibr" rid="b48">48</xref><xref ref-type="bibr" rid="b49">49</xref>; therefore, we measured free SA and glucosyl-SA levels in the leaves of niclosamide-treated rice plants. SA levels increased in response to niclosamide treatment (<xref ref-type="fig" rid="f7">Fig. 7a,b</xref>), indicating that niclosamide induces the expression of pathogenesis-related genes by increasing the levels of SA and SA-conjugates.</p></sec><sec disp-level="2"><title>Niclosamide has no effect on rice growth and development</title><p>We examined the effect of niclosamide on rice growth and development from the vegetative stage to seed maturation. Three-week-old rice plants were treated with 8 μg/ml niclosamide at 4-day intervals and the phenotypic characteristics of niclosamide-treated and -untreated plants were examined at 80 days after planting. Plant height and leaf characteristics were not altered by niclosamide treatment, although the contents of chlorophyll and other compounds (SPAD values) were slightly reduced in niclosamide-treated plants (<xref ref-type="fig" rid="f8">Fig. 8a–g</xref>). Seed characteristics such as color, number, and weight were not affected by niclosamide treatment (<xref ref-type="fig" rid="f8">Fig. 8h–l</xref>), indicating that niclosamide does not have any detrimental effects on rice growth, development, and grain yield.</p></sec></sec><sec disp-level="1"><title>Discussion</title><p><italic>Xoo</italic>-mediated leaf blight is one of the most devastating rice diseases worldwide. For the past two decades, substantial efforts have been made to identify and isolate bacterial blight-resistance genes. In this study, we used an alternative chemical approach to protect rice from <italic>Xoo</italic> infection.</p><p>More than 30 drugs, including antibiotics, have been utilized to protect crops from pathogen attack. Some of these have broad-spectrum bactericidal and fungicidal activity, whereas others specifically target bacteria or fungi. Oxytetracycline and streptomycin are commonly used antibiotics in humans and plants<xref ref-type="bibr" rid="b15">15</xref>, suggesting that some types of human drugs can positively control diseases in plants. Therefore, we screened human drugs for their ability to prevent rice leaf blight disease. Our long-term goal is to identify a master regulator that inhibits pathogenic disease in both plants and humans.</p><p>Niclosamide was initially characterized as an oral antihelminthic drug and molluscicide<xref ref-type="bibr" rid="b36">36</xref>. Several recent studies report that niclosamide is active against cancer cells<xref ref-type="bibr" rid="b39">39</xref><xref ref-type="bibr" rid="b50">50</xref><xref ref-type="bibr" rid="b51">51</xref>, which indicates that niclosamide has broad-spectrum disease control activity. Therefore, we examined whether niclosamide had inhibitory activity against rice leaf blight, and found that the leaf blight symptoms were suppressed by niclosamide through inhibition of <italic>Xoo</italic> growth and lesion development (<xref ref-type="fig" rid="f2">Figs 2</xref> and <xref ref-type="fig" rid="f3">3</xref>). Niclosamide moved systemically from the local application site to <italic>Xoo</italic>-inoculated distal leaves, and inhibited lesion development and leaf wilting by inhibiting <italic>Xoo</italic> growth in distal leaves (<xref ref-type="fig" rid="f4">Fig. 4</xref>). We examined the effect of niclosamide on the growth of three <italic>Xoo</italic> strains, and found that it inhibited their growth (<xref ref-type="fig" rid="f1">Fig. 1a,e</xref>). Next, we evaluated whether niclosamide induced the expression of the defense-related genes <italic>OsPR1</italic>, <italic>OsPR10</italic>, <italic>OsWRKY45</italic>, <italic>OsWRKY62</italic>, <italic>OsWRKY71</italic>, and <italic>OsHI-LOX</italic>, and detected higher transcript levels for these genes (<xref ref-type="fig" rid="f6">Fig. 6</xref>). These results clearly indicate that niclosamide blocks the development of rice leaf blight by directly inhibiting <italic>Xoo</italic> bacterial growth (<xref ref-type="fig" rid="f2">Figs 2</xref>c and <xref ref-type="fig" rid="f3">3</xref>c) and/or by inducing defense-related gene expression (<xref ref-type="fig" rid="f6">Fig. 6</xref>). Niclosamide also inhibited <italic>Xag</italic> growth, which causes bacterial leaf pustule disease in soybean<xref ref-type="bibr" rid="b45">45</xref>, although its inhibitory effect against <italic>Xag</italic> was weaker than that against <italic>Xoo</italic> strains (<xref ref-type="fig" rid="f1">Fig. 1a</xref>). This suggests that niclosamide may inhibit the growth of select bacterial pathogens of other crops and thereby protect them from disease.</p><p>Jasmonate (JA) and SA function in plant defense pathways by inducing the expression of numerous genes<xref ref-type="bibr" rid="b52">52</xref>. Treatment of rice leaves with benzothiadiazole or probenazole induced SA accumulation and increased resistance to bacterial blight and fungal blast caused by <italic>Xoo</italic> and <italic>M. grisea</italic>, respectively<xref ref-type="bibr" rid="b13">13</xref><xref ref-type="bibr" rid="b14">14</xref><xref ref-type="bibr" rid="b34">34</xref>. Our qRT-PCR analysis showed that niclosamide induced the expression of SA-dependent genes in rice leaves (<xref ref-type="fig" rid="f6">Fig. 6</xref>). We investigated whether niclosamide induced defense mechanisms directly by functioning like a phytohormone, or indirectly via SA-mediated signaling pathways. We measured the levels of free SA and its conjugate glucosyl-SA, and found that their levels were higher in niclosamide-treated leaves than in control leaves (<xref ref-type="fig" rid="f7">Fig. 7</xref>). This indicates that increase in SA level can contribute to the blockage of <italic>Xoo</italic> lesion development by niclosamide.</p><p>WRKY proteins are involved in JA response pathways in rice. OsWRKY30 triggers the expression of JA-responsive genes and JA accumulation, and provides rice with resistance to fungal pathogens <italic>Rhizoctonia solani</italic> and <italic>M. grisea</italic><xref ref-type="bibr" rid="b33">33</xref>. <italic>OsWRKY45</italic> expression regulates JA accumulation and resistance to the rice blast fungus <italic>M. grisea</italic><xref ref-type="bibr" rid="b53">53</xref>. Our results showed that the expression of <italic>OsHI-LOX</italic>, a JA-responsive gene, was induced in niclosamide-treated leaves (<xref ref-type="fig" rid="f6">Fig. 6</xref>). This indicates that JA-dependent defense signaling pathways can also contribute to the blockage of <italic>Xoo</italic> lesion development by niclosamide.</p><p>Pathogens are sensitive to the activities of certain classes of drugs. The antibiotic streptomycin, which is commonly used in animals and humans, protects crops from some bacterial and fungal pathogens<xref ref-type="bibr" rid="b15">15</xref>. Niclosamide serves as an oral antihelminthic drug, molluscicide, and anti-tumor agent in human disease; therefore, we reasoned that niclosamide might have beneficial effects on bacterial and fungal diseases of rice. Niclosamide inhibited growth of the fungal pathogen <italic>M. oryzae</italic>, although this effect was much weaker than that against pathogenic <italic>Xoo</italic> bacteria (<xref ref-type="fig" rid="f1">Fig. 1b</xref>). This suggests that niclosamide may inhibit the growth of other fungal pathogens and thereby protect plants from fungal diseases. Benzothiadiazole and probenazole induce SA pathway-mediated defense responses in plants by acting as chemical inducers<xref ref-type="bibr" rid="b13">13</xref><xref ref-type="bibr" rid="b14">14</xref><xref ref-type="bibr" rid="b34">34</xref><xref ref-type="bibr" rid="b54">54</xref><xref ref-type="bibr" rid="b55">55</xref><xref ref-type="bibr" rid="b56">56</xref><xref ref-type="bibr" rid="b57">57</xref>, leading to strong resistance against bacterial and fungal pathogens. Niclosamide induces the expression of both SA- and JA-responsive genes in rice (<xref ref-type="fig" rid="f6">Fig. 6</xref>). This indicates that niclosamide may also be effective against fungi due to JA. Further study of the effects of niclosamide against other fungal pathogens will indicate whether the compound can protect plants from diseases caused by fungal pathogens.</p><p>The biochemical function and physiological action of niclosamide in human health and disease have been elucidated. Niclosamide inhibits glucose uptake, mitochondrial oxidative phosphorylation, and anaerobic metabolism in parasitic helminths<xref ref-type="bibr" rid="b44">44</xref>. It also inhibits transcription and DNA binding of the NFκΒ pathway, and increases ROS levels to induce apoptosis in acute myelogenous leukemia cells<xref ref-type="bibr" rid="b50">50</xref>. Recent work reports that niclosamide inhibits <italic>Pseudomonas aeruginosa</italic> quorum sensing<xref ref-type="bibr" rid="b58">58</xref>. These results suggest that niclosamide controls signaling networks of prokaryotes and eukaryotes by modulating metabolic and signaling pathways. Currently, we do not know the mechanism underlying niclosamide-mediated inhibition of <italic>Xoo</italic> growth and induction of SA-and JA-dependent plant defense responses. Future work will examine the effects of niclosamide on glucose metabolism, the transcriptome and proteome, and quorum sensing to identify the mechanism by which niclosamide inhibits bacterial leaf blight in rice. Further investigations of the effects of niclosamide on mitochondrial oxidative phosphorylation, changes in transcriptome and ROS levels, and changes in hormone signals (including SA and JA) also are required to fully characterize its functions as a broad-spectrum signaling regulator in plants.</p><p>The use of antibiotics and chemicals in plant agriculture is a subject of some concern. The use of antibiotics and chemicals in open fields over large expanses of land may increase the frequency of antibiotic resistance in gene pools, or possible chemical accumulation in plant tissues and the food chain. Niclosamide is a teniacide, which is effective against cestodes such as tapeworms that infect humans and many other animals (although it is not effective against as pinworms and roundworms). Niclosamide also is used as a piscicide. Recent work reported that it appeared to be safe and well tolerated in humans, even when used in mass treatment campaigns in several countries, although it has known adverse effects including nausea, retching, abdominal pain, light headedness, pruritus, vomiting, and dizziness<xref ref-type="bibr" rid="b59">59</xref>. There are no previous reports of niclosamide effects on plant growth, other organisms, or the food chain. We cannot state that niclosamide treatment of agricultural crops is safe for other organisms or for the food chain because research in this area is still lacking. Therefore, further studies on the function and stability of niclosamide under natural environmental conditions and when applied to crop plants are required to determine whether niclosamide has adverse effects on other organisms and human health when applied to plants.</p><p>In conclusion, the results presented herein indicate that niclosamide protects rice plants from bacterial leaf blight by inhibiting <italic>Xoo</italic> growth, inducing SA accumulation, and/or by inducing the expression of defense-related gene pathways. Further functional studies on niclosamide-mediated growth inhibition of bacterial and fungal pathogens, and elucidation of its role in plant signaling pathways, will provide information about the mechanism underlying niclosamide activity in plants. Field tests evaluating the effects of niclosamide application on rice plants will provide information on whether it has adverse effects on other organisms or the food chain.</p></sec><sec disp-level="1"><title>Methods</title><sec disp-level="2"><title>Plant materials and chemicals</title><p>The rice (<italic>Oryza sativa</italic>) japonica cultivar, Nipponbare, was used in this study. Before treatment with the pathogen or niclosamide, plants were grown in a greenhouse maintained at 31 °C in the light and 25 °C in the dark under a 16 h photoperiod until the booting stage. The chamber was equipped with incandescent lights. Stock solutions of niclosamide (Sigma-Aldrich) and parthenolide (Sigma-Aldrich) were made by dissolving the chemicals in dimethyl sulfoxide (DMSO) at a concentration of 5 mg/ml. The stock solutions were diluted in distilled water before use.</p></sec><sec disp-level="2"><title>Examining the effect of niclosamide on <italic>Xoo</italic> bacterial growth</title><p>The effect of niclosamide on the growth of <italic>Xoo</italic> strains PXO99, 10208, and K3a, and a <italic>Xag</italic> strain, and the MIC of niclosamide against the PXO99 and 10208 strains were investigated. <italic>Xoo</italic> and <italic>Xag</italic> strains were obtained from the Rural Development Administration, Korea. Bacterial strains were cultivated in peptone-sucrose broth containing 15 μg/ml of the antibiotic cephalexin (Sigma-Aldrich), and grown to an optical density of 1.0 at 600 nm. Then, 2 μl of each culture was grown on agar medium containing different concentrations of niclosamide (0−50 μg/ml) to examine the effects of niclosamide on bacterial growth. Inoculated plates were incubated at 28 °C for 48 h and then photographed. The same method was used to determine whether parthenolide had bactericidal activity against the PXO99 and 10208 strains. The bactericidal activity of niclosamide on three different <italic>E. coli</italic> strains, Top10, Rosseta2, and DH10b, also was examined. The <italic>E. coli</italic> strains were cultivated in LB medium to an optical density of 1.0 at 600 nm. Then, 2 μl of each culture was grown on agar medium containing different concentrations of niclosamide. Inoculated plates were incubated at 37 °C for 24 h and photographed.</p><p>To determine the MIC, 2 μl of each culture were inoculated onto peptone-sucrose agar (PSA) medium containing 0−5.0 μg/ml niclosamide. Bacterial growth was not detected at a concentration of 5.0 μg/ml niclosamide. Thus, to determine a more specific MIC value, the MIC of niclosamide against both <italic>Xoo</italic> strains was narrowed down from 4.0 to 4.8 μg/ml. The lowest concentration of niclosamide (4.2 μg/ml) with no visible growth was taken as the MIC.</p><p>To examine the effects of niclosamide on fungal growth, 12 <italic>M. oryzae</italic> strains (46531, 46532, 46534, 46535, 46536, 46538, 46540, 46541, 46542, 46544, KI215, and PO6-6) were inoculated onto potato dextrose agar (Difco) medium containing different concentrations of niclosamide (0−50 μg/ml). The plates were then incubated at 28 °C for 72 h and photographed.</p></sec><sec disp-level="2"><title>Effect of niclosamide dosage on rice disease responses</title><p>To determine the minimum amount of niclosamide needed to inhibit the development of leaf blight in rice, the rice cultivar Nipponbare and <italic>Xoo</italic> strain PXO99 were used. PXO99 cells were prepared as follows. A single PXO99 colony was suspended in peptone-sucrose broth and plated onto fresh PSA medium. After 2 days of culture at 28 °C, the cells were collected and centrifuged to remove exopolysaccharides. The cells were then resuspended in double-distilled water, producing more than 10<sup>9</sup> cell forming units (cfu) per ml.</p><p>The dosage effect of niclosamide on PXO99 lesion development was evaluated by measuring the lesion length (cm) and bacterial growth in the leaves of niclosamide-treated rice. To treat rice with PXO99, plants were grown for 80 days in a greenhouse at 31 °C in the light and 25 °C in the dark under a 16 h photoperiod until the booting stage. The greenhouse humidity was maintained over 90%. The fully expanded uppermost leaves were inoculated with PXO99 by the leaf-clipping method, followed by treatment with various doses (0, 4, 8, 20, and 40 μg/ml) of niclosamide applied by foliar spraying every 4 days. As a control, mock inoculation was conducted with distilled water. After 16 days, the leaves were photographed and lesion length was measured using ImageJ Software. All data are expressed as the mean values from 15 leaves per treatment. The experiment was repeated five times under the same conditions. To estimate bacterial growth, leaves were cut off the plants and ground with a mortar and pestle. After grinding, each sample was suspended in sterile water and plated onto PSA medium containing 15 μg/ml cephalexin. The bacterial population was scored by counting the number of colonies every 4 days after inoculation. The lesion lengths and bacterial populations were expressed as the mean value plus/minus the standard deviation.</p></sec><sec disp-level="2"><title>Niclosamide effects on disease responses in rice</title><p>Rice plants were inoculated with <italic>Xoo</italic> and sprayed with niclosamide. PXO99 cells and plants were prepared as described above. To treat rice with PXO99, the fully expanded uppermost leaves of 80-day-old rice plants were inoculated with PXO99 by the leaf-clipping method. The samples were incubated for 4 days until leaf blight lesions with a length of 2 cm developed, after which the leaves were treated with 8 μg/ml niclosamide by foliar spraying every 4 days. As a control, mock inoculation was conducted with distilled water. Leaves were photographed after 16 days, and lesion length and bacterial growth estimated as described above. All data are expressed as the mean values of 15 leaves per treatment. The experiment was repeated five times under the same conditions. The lesion length and bacterial population were expressed as the mean value plus/minus the standard deviation.</p></sec><sec disp-level="2"><title>Systemic effect of niclosamide on rice disease responses</title><p>To examine the systemic effect of niclosamide on <italic>Xoo</italic>-mediated leaf blight development, the fully expanded uppermost leaves of 80-day-old rice plants were inoculated with PXO99 by the leaf-clipping as described above. Half of the leaves were completely covered with polythene bags and half were sprayed with 8 μg/ml niclosamide. The local leaves were then sprayed with 8 μg/ml niclosamide every 4 days. Both local and systemic leaves were photographed after 16 days. Lesion length and bacterial growth were estimated as described above. The experiment was repeated five times under the same conditions. The lesion length and bacterial population were expressed as the mean value plus/minus the standard deviation.</p></sec><sec disp-level="2"><title>Quantification of systemically translocated niclosamide</title><p>Half of the leaves of 80-day-old rice plants were covered by polythene bags and the remaining systemic leaves were sprayed with 8 μg/ml niclosamide. After treatment, the bag-covered leaves were harvested after incubation for the indicated time periods. Niclosamide was extracted from 0.5 g of each sample using absolute MeOH, and the niclosamide concentration was determined by HPLC separation and fluorescence detection. The relative niclosamide concentration in each sample was compared with the 100 ppb niclosamide standard. Niclosamide levels were expressed as the mean value plus/minus the standard deviation.</p></sec><sec disp-level="2"><title>Determination of salicylic acid (SA) and its conjugate</title><p>SA and the SA conjugate glucosyl-SA were extracted from 0.5 g of rice leaves that were treated with 8 μg/ml niclosamide for 4 h. Samples were collected at five time points (0, 0.5, 1, 2, and 4 h). The concentrations of SA and glucosyl-SA were measured by HPLC as described previously<xref ref-type="bibr" rid="b60">60</xref>.</p></sec><sec disp-level="2"><title>Estimation of pathogen-related gene transcript levels in niclosamide-treated rice</title><p>The rice cultivar Nipponbare was sprayed with 40 μg/ml niclosamide as described above. Total RNA was extracted from niclosamide-treated and untreated leaves at 0, 0.5, 12, and 24 h, quantified, and diluted to equal concentrations. All niclosamide-treated or untreated plants were used for total RNA extraction. First-strand cDNA was synthesized using 1 μg of total RNA and an iScript cDNA Synthesis Kit (Bio-Rad). An equal volume of cDNA was amplified by quantitative real-time RT-PCR (MyiQ, Bio-Rad) according to the manufacturer’s protocol. Then, 10 nM of specific primers and template cDNA were combined with 25 μl iQ SYBR Green Super Mix (Bio-Rad) and the amplification reaction performed under the following thermal cycling conditions: 95 °C for 10 min; 45 cycles of 95 °C for 10 s; 60 °C for 10 s; and 72 °C for 10 s. The CT values of target genes were normalized to the CT value of the <italic>actin1</italic> gene and analyzed with iCycler IQ software (Bio-Rad). The experiments were repeated three times. PCR primers were designed using Primer3Plus (<ext-link ext-link-type="uri" xlink:href="http://www.bioinformatics.nl/cgi-bin/primer3plus/primer3plus.cgi/">http://www.bioinformatics.nl/cgi-bin/primer3plus/primer3plus.cgi/</ext-link>). Primer specificity was verified by cloning into the pGEM T-Easy vector (Promega) and sequencing with an ABI 3730 × l DNA Analyzer (Applied Biosystems). The primers used for quantitative PCR were as follows: <italic>OsPR1</italic> forward, 5′-TTATCCTGCTGCTTGCTGGT-3′; <italic>OsPR1</italic> reverse, 5′-GATGTTCTCGCCGTACTTCC-3′; <italic>OsPR10</italic> forward, 5′-GGCACCATCTACACCATGAA-3′ and <italic>OsPR10</italic> reverse, 5′-TTGTCGGCTGTGATGAATGT-3′; <italic>OsWRKY45</italic> forward, 5′-CCGGCATGGAGTTCTTCAAG-3′ and <italic>OsWRKY45</italic> reverse, 5′-TATTTCTGTACACACGCGTGGAA-3′; <italic>OsWRKY62</italic> forward, 5′-AGGATGGGTACCAATGGA-3′ and <italic>OsWRKY62</italic> reverse, 5′-ACGAGTTGATGGAGATGGA-3′; <italic>OsWRKY71</italic> forward, 5′-AGCCCAAGATCTCCAAGCTC-3′ and <italic>OsWRKY71</italic> reverse, 5′-ACGAGGATCGTGTTGTCCTC-3′; <italic>OsHI-LOX</italic> forward, 5′-GCATCCCCAACAGCACATC-3′ and <italic>OsHI-LOX</italic> reverse, 5′-AATAAAGATTTGGGAGTGACATATTGG-3′; <italic>OsACS2</italic> forward, 5′-GGAATAAAGCTGCTGCCGAT-3′ and <italic>OsACS2</italic> reverse, 5′-TGAGCCTGAAGTCGTTGAAGC-3′.</p></sec><sec disp-level="2"><title>Niclosamide effect on rice growth and development</title><p>Three-week-old Nipponbare leaves were sprayed with 8 μg/ml niclosamide at 4-day intervals until seed maturation. The phenotypic characteristics of untreated and niclosamide-treated plants were measured at 70 days after planting. Grain phenotypes were examined after seed maturation.</p></sec></sec><sec disp-level="1"><title>Additional Information</title><p><bold>How to cite this article</bold>: Kim, S.-I. <italic>et al.</italic> Niclosamide inhibits leaf blight caused by <italic>Xanthomonas oryzae</italic> in rice. <italic>Sci. Rep.</italic><bold>6</bold>, 21209; doi: 10.1038/srep21209 (2016).</p></sec></body><back><ack><p>We are grateful to Dr. Hong-Gu Kang in Texas State University for critical reading of the manuscript. This work was supported by a grant from the Next-Generation BioGreen 21 Program (Plant Molecular Breeding Center no. PJ01108701), Rural Development Administration, Republic of Korea.</p></ack><ref-list><ref id="b1"><mixed-citation publication-type="journal"><name><surname>Mew</surname><given-names>T. W.</given-names></name><article-title>Current status and future prospects of research on bacterial blight of rice</article-title>. <source>Annu. Rev. Phytopathol.</source><volume>25</volume>, <fpage>359</fpage>–<lpage>382</lpage> (<year>1987</year>).</mixed-citation></ref><ref id="b2"><mixed-citation publication-type="journal"><name><surname>Tabei</surname><given-names>H.</given-names></name><article-title>Anatomical studies of rice plant affected with bacterial leaf blight, <italic>Xanthomonas oryzae</italic> (Uyeda et Ishiyama Dowson)</article-title>. <source>Bull. Kyushu Agri. Expt. Sta</source>. <volume>19</volume>, <fpage>193</fpage>–<lpage>257</lpage> (<year>1977</year>).</mixed-citation></ref><ref id="b3"><mixed-citation publication-type="journal"><name><surname>Ishiyama</surname><given-names>S.</given-names></name><article-title>Studies of Bacterial Leaf Blight of Rice</article-title>. <source>Report Imp. Agric. Stn. Konosu</source>. <volume>45</volume>, <fpage>233</fpage>–<lpage>261</lpage> (<year>1922</year>).</mixed-citation></ref><ref id="b4"><mixed-citation publication-type="journal"><name><surname>Ray</surname><given-names>S. K.</given-names></name>, <name><surname>Rajeshwari</surname><given-names>R.</given-names></name> & <name><surname>Sonti</surname><given-names>R. V.</given-names></name><article-title>Mutants of <italic>Xanthomonas oryzae</italic> pv. <italic>oryzae</italic> deficient in general secretory pathway are virulence deficient and unable to secrete xylanase</article-title>. <source>Mol. Plant-Microbe Interact</source>. <volume>13</volume>, <fpage>394</fpage>–<lpage>401</lpage> (<year>2000</year>).<pub-id pub-id-type="pmid">10755302</pub-id></mixed-citation></ref><ref id="b5"><mixed-citation publication-type="journal"><name><surname>Köplin</surname><given-names>R.</given-names></name><italic>et al.</italic><article-title>Genetics of xanthan production in <italic>Xanthomonas campestris: the xanA</italic> and <italic>xanB</italic> gene are involved in UDP-glucose and GDP-mannose biosynthesis</article-title>. <source>J. Bacteriol.</source><volume>174</volume>, <fpage>191</fpage>–<lpage>199</lpage> (<year>1992</year>).<pub-id pub-id-type="pmid">1370280</pub-id></mixed-citation></ref><ref id="b6"><mixed-citation publication-type="journal"><name><surname>Hu</surname><given-names>J.</given-names></name>, <name><surname>Qian</surname><given-names>W.</given-names></name> & <name><surname>He</surname><given-names>C.</given-names></name><article-title>The <italic>Xanthomonas oryzae</italic> pv. <italic>oryzae eglXoB</italic> endoglucanase gene is required for virulence to rice</article-title>. <source>FEMS Microbiol. Lett.</source><volume>269</volume>, <fpage>273</fpage>–<lpage>279</lpage> (<year>2007</year>).<pub-id pub-id-type="pmid">17326805</pub-id></mixed-citation></ref><ref id="b7"><mixed-citation publication-type="journal"><name><surname>Rajeshwari</surname><given-names>R.</given-names></name>, <name><surname>Jha</surname><given-names>G.</given-names></name> & <name><surname>Sonti</surname><given-names>R. V.</given-names></name><article-title>Role of an <italic>in planta</italic> expressed xylanase of <italic>Xanthomonas oryzae</italic> pv. <italic>oryzae</italic> in promoting virulence on rice</article-title>. <source>Mol. Plant-Microbe Interact</source>. <volume>18</volume>, <fpage>830</fpage>–<lpage>837</lpage> (<year>2005</year>).<pub-id pub-id-type="pmid">16134895</pub-id></mixed-citation></ref><ref id="b8"><mixed-citation publication-type="journal"><name><surname>Jha</surname><given-names>G.</given-names></name>, <name><surname>Rajeshwari</surname><given-names>R.</given-names></name> & <name><surname>Sonti</surname><given-names>R. V.</given-names></name><article-title>Functional interplay between two <italic>Xanthomonas oryzae</italic> pv. <italic>oryzae</italic> secretion systems in modulating virulence on rice</article-title>. <source>Mol. Plant-Microbe Interact</source>. <volume>20</volume>, <fpage>31</fpage>–<lpage>40</lpage> (<year>2007</year>).<pub-id pub-id-type="pmid">17249420</pub-id></mixed-citation></ref><ref id="b9"><mixed-citation publication-type="journal"><name><surname>Chithrashree</surname><given-names>A. C.</given-names></name>, <name><surname>Udayashankar</surname><given-names>S.</given-names></name>, <name><surname>Chandra Nayaka</surname><given-names>M. S.</given-names></name> & <name><surname>Srinivas</surname><given-names>R. C.</given-names></name><article-title>Plant growth-promoting rhizobacteria mediate induced systemic resistance in rice against bacterial leaf blight caused by <italic>Xanthomonas oryzae</italic> pv<italic>. Oryzae</italic></article-title>. <source>Biol. Control.</source><volume>59</volume>, <fpage>114</fpage>–<lpage>122</lpage> (<year>2011</year>).</mixed-citation></ref><ref id="b10"><mixed-citation publication-type="journal"><name><surname>Gnanamanickam</surname><given-names>S. S.</given-names></name><source>Biological control of rice diseases</source>. New York: Martial Decker <volume>Vol. 8</volume> Ch. 5, <fpage>67</fpage>–<lpage>78</lpage> (Springer, <year>2002)</year>.</mixed-citation></ref><ref id="b11"><mixed-citation publication-type="journal"><name><surname>Chand</surname><given-names>T.</given-names></name>, <name><surname>Sing</surname><given-names>N.</given-names></name>, <name><surname>Sing</surname><given-names>H.</given-names></name> & <name><surname>Thind</surname><given-names>B. S.</given-names></name><article-title>Field efficacy of stable bleaching powder to control bacterial blight of rice in rice</article-title>. <source>Int. Rice Res. New</source>. <volume>4</volume>, <fpage>12</fpage>–<lpage>13</lpage> (<year>1979</year>).</mixed-citation></ref><ref id="b12"><mixed-citation publication-type="journal"><name><surname>Khan</surname><given-names>J. A.</given-names></name><italic>et al.</italic><article-title>Chemical control of bacterial leaf blight of rice caused by <italic>Xanthomonas oryzae</italic> pv. <italic>Oryzae. Pakitsan</italic></article-title><source>J. Phytopathol.</source><volume>24</volume>, <fpage>97</fpage>–<lpage>100</lpage> (<year>2012</year>).</mixed-citation></ref><ref id="b13"><mixed-citation publication-type="journal"><name><surname>Karthikeyan</surname><given-names>V.</given-names></name> & <name><surname>Gnanamanickam</surname><given-names>S. S.</given-names></name><article-title>Induction of systemic resistance in rice to bacterial blight by 1,2,3-benzothiadiazole 7-carbothioic acid-S-methyl ester (BTH) treatments</article-title>. <source>Arch. Phytopathol. Plant Protect</source>. <volume>44</volume>, <fpage>269</fpage>–<lpage>281</lpage> (<year>2011</year>).</mixed-citation></ref><ref id="b14"><mixed-citation publication-type="journal"><name><surname>Iwai</surname><given-names>T.</given-names></name>, <name><surname>Seo</surname><given-names>S.</given-names></name>, <name><surname>Mitsuhara</surname><given-names>I.</given-names></name> & <name><surname>Ohashi</surname><given-names>Y.</given-names></name><article-title>Probenazole-induced accumulation of salicylic acid confers resistance to <italic>Magnaporthe grisea</italic> in adult rice plants</article-title>. <source>Plant Cell Physiol.</source><volume>48</volume>, <fpage>915</fpage>–<lpage>924</lpage> (<year>2007</year>).<pub-id pub-id-type="pmid">17517758</pub-id></mixed-citation></ref><ref id="b15"><mixed-citation publication-type="journal"><name><surname>McManus</surname><given-names>P. S.</given-names></name>, <name><surname>Stockwell</surname><given-names>V. O.</given-names></name>, <name><surname>Sundin</surname><given-names>G. W.</given-names></name> & <name><surname>Jones</surname><given-names>A. L.</given-names></name><article-title>Antibiotic use in plant agriculture</article-title>. <source>Annu. Rev. Phytopathol.</source><volume>40</volume>, <fpage>443</fpage>–<lpage>465</lpage> (<year>2002</year>).<pub-id pub-id-type="pmid">12147767</pub-id></mixed-citation></ref><ref id="b16"><mixed-citation publication-type="journal"><name><surname>Bhasin</surname><given-names>H.</given-names></name><italic>et al.</italic><article-title>New PCR-based sequence-tagged site marker for bacterial blight resistance gene <italic>Xa38</italic> of rice</article-title>. <source>Mol. Breeding.</source><volume>30</volume>, <fpage>607</fpage>–<lpage>611</lpage> (<year>2012</year>).</mixed-citation></ref><ref id="b17"><mixed-citation publication-type="journal"><name><surname>Natrajkumar</surname><given-names>P.</given-names></name><italic>et al.</italic><article-title>Identification and fine-mapping of <italic>Xa33</italic>, a novel gene for resistance to <italic>Xanthomonas oryzae</italic> pv. <italic>oryzae</italic></article-title>. <source>Phytopathol</source>. <volume>102</volume>, <fpage>222</fpage>–<lpage>228</lpage> (<year>2012</year>).</mixed-citation></ref><ref id="b18"><mixed-citation publication-type="journal"><name><surname>Song</surname><given-names>W. Y.</given-names></name><italic>et al.</italic><article-title>Evolution of the rice <italic>Xa21</italic> disease resistance gene family</article-title>. <source>Plant Cell</source><volume>9</volume>, <fpage>1279</fpage>–<lpage>1287</lpage> (<year>1997</year>).<pub-id pub-id-type="pmid">9286106</pub-id></mixed-citation></ref><ref id="b19"><mixed-citation publication-type="journal"><name><surname>Yang</surname><given-names>D.</given-names></name><italic>et al.</italic><article-title>Construction of a BAC contig containing the <italic>xa5</italic> locus in rice</article-title>. <source>Theor. Appl. Genet.</source><volume>97</volume>, <fpage>1120</fpage>–<lpage>1124</lpage> (<year>1998</year>).</mixed-citation></ref><ref id="b20"><mixed-citation publication-type="journal"><name><surname>Gu</surname><given-names>K. Y.</given-names></name><italic>et al.</italic><article-title>R gene expression induced by a type-III effectors triggers disease resistance in rice</article-title>. <source>Nature</source><volume>435</volume>, <fpage>1122</fpage>–<lpage>1125</lpage> (<year>2005</year>).<pub-id pub-id-type="pmid">15973413</pub-id></mixed-citation></ref><ref id="b21"><mixed-citation publication-type="journal"><name><surname>Liu</surname><given-names>D. O.</given-names></name>, <name><surname>Ronald</surname><given-names>P. C.</given-names></name> & <name><surname>Bogdanove</surname><given-names>A. J.</given-names></name><article-title><italic>Xanthomonas oryzae pathovars</italic>: model pathogens of a model crop</article-title>. <source>Mol. Plant Pathol.</source><volume>7</volume>, <fpage>303</fpage>–<lpage>324</lpage> (<year>2006</year>).<pub-id pub-id-type="pmid">20507449</pub-id></mixed-citation></ref><ref id="b22"><mixed-citation publication-type="journal"><name><surname>Cheema</surname><given-names>K. K.</given-names></name><italic>et al.</italic><article-title>A novel bacterial blight resistance gene from <italic>Oryza nivara</italic> mapped to 38 kb region on chromosome 4 L and transferred to <italic>Oryza sativa</italic> L</article-title>. <source>Genet. Res.</source><volume>90</volume>, <fpage>397</fpage>–<lpage>407</lpage> (<year>2008</year>).</mixed-citation></ref><ref id="b23"><mixed-citation publication-type="journal"><name><surname>Yoshimura</surname><given-names>S.</given-names></name><italic>et al.</italic><article-title>Expression of <italic>Xa1</italic>, a bacterial blight resistance gene in rice, is induced by bacterial inoculation</article-title>. <source>Proc. Natl. Acad. Sci. USA</source><volume>95</volume>, <fpage>1663</fpage>–<lpage>1668</lpage> (<year>1998</year>).<pub-id pub-id-type="pmid">9465073</pub-id></mixed-citation></ref><ref id="b24"><mixed-citation publication-type="journal"><name><surname>Hulbert</surname><given-names>S. H.</given-names></name>, <name><surname>Webb</surname><given-names>C. A.</given-names></name>, <name><surname>Smith</surname><given-names>S. M.</given-names></name> & <name><surname>Sun</surname><given-names>Q.</given-names></name><article-title>Resistance genes complexes: evolution and utilization.</article-title><source>Ann. Rev. Phytopathol.</source><volume>39</volume>, <fpage>258</fpage>–<lpage>312</lpage> (<year>2011</year>).</mixed-citation></ref><ref id="b25"><mixed-citation publication-type="journal"><name><surname>White</surname><given-names>F. F.</given-names></name> & <name><surname>Yang</surname><given-names>B.</given-names></name><article-title>Host and pathogen factors controlling the rice-<italic>Xanthomonas oryzae</italic> interaction</article-title>. <source>Plant Physiol.</source><volume>150</volume>, <fpage>1677</fpage>–<lpage>1686</lpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19458115</pub-id></mixed-citation></ref><ref id="b26"><mixed-citation publication-type="journal"><name><surname>Verdier</surname><given-names>V.</given-names></name>, <name><surname>Cruz</surname><given-names>C. V.</given-names></name> & <name><surname>Leach</surname><given-names>J. E.</given-names></name><article-title>Controlling rice bacterial blight in africa: needs and prospects</article-title>. <source>J. Biotechnol.</source><volume>159</volume>, <fpage>320</fpage>–<lpage>328</lpage> (<year>2012</year>).<pub-id pub-id-type="pmid">21963588</pub-id></mixed-citation></ref><ref id="b27"><mixed-citation publication-type="journal"><name><surname>Singh</surname><given-names>S.</given-names></name><italic>et al.</italic><article-title>Pyramiding three bacterial blight resistance genes (<italic>xa5</italic>, <italic>xa13</italic> and <italic>Xa21</italic>) using marker-assisted selection into indica rice cultivar PR106</article-title>. <source>Theor. Appl.Genet.</source><volume>102</volume>, <fpage>1011</fpage>–<lpage>1015</lpage> (<year>2012</year>).</mixed-citation></ref><ref id="b28"><mixed-citation publication-type="journal"><name><surname>Lia</surname><given-names>Q.</given-names></name><italic>et al.</italic><article-title>Expression profiling of rice genes in early defense responses to blast and bacterial blight pathogens using cDNA microarray</article-title>. <source>Physiol. Mol. Plant Pathol.</source><volume>68</volume>, <fpage>51</fpage>–<lpage>60</lpage> (<year>2006</year>).</mixed-citation></ref><ref id="b29"><mixed-citation publication-type="journal"><name><surname>Cernadas</surname><given-names>R. A.</given-names></name><italic>et al.</italic><article-title>Code-assisted discovery of TAL Effector targets in bacterial leaf streak of rice reveals contrast with bacterial blight and a novel susceptibility gene</article-title>. <source>PLoS Pathog.</source><volume>10</volume>, <fpage>e1003972</fpage> (<year>2014</year>).<pub-id pub-id-type="pmid">24586171</pub-id></mixed-citation></ref><ref id="b30"><mixed-citation publication-type="journal"><name><surname>Sharma</surname><given-names>A.</given-names></name><italic>et al.</italic><article-title>Transgenic expression of cecropin B, an antibacterial peptide from <italic>Bombyx mori</italic>, confers enhaced restance to bacterial leaf blight in rice</article-title>. <source>FEBS lett.</source><volume>484</volume>, <fpage>7</fpage>–<lpage>11</lpage> (<year>2000</year>).<pub-id pub-id-type="pmid">11056212</pub-id></mixed-citation></ref><ref id="b31"><mixed-citation publication-type="journal"><name><surname>Swamy</surname><given-names>P.</given-names></name><italic>et al.</italic><article-title>Evaluation of bacterial blight resistance in rice lines carrying multiple resistance genes and Xa21 transgenic lines</article-title>. <source>Curr. Sci.</source><volume>90</volume>, <fpage>818</fpage>–<lpage>824</lpage> (<year>2006</year>).</mixed-citation></ref><ref id="b32"><mixed-citation publication-type="journal"><name><surname>Han</surname><given-names>M.</given-names></name><italic>et al.</italic><article-title>OsWRKY30 is a transcriptional activator that enhances rice resistance to the <italic>Xanthomonas oryzae pathovar oryzae</italic></article-title>. <source>J. Plant Biol.</source><volume>56</volume>, <fpage>258</fpage>–<lpage>265</lpage> (<year>2013</year>).</mixed-citation></ref><ref id="b33"><mixed-citation publication-type="journal"><name><surname>Peng</surname><given-names>X.</given-names></name><italic>et al.</italic><article-title>Constitutive expression of rice <italic>WRKY30</italic> gene increases the endogenous jasmonic acid accumulation, <italic>PR</italic> gene expression and resistance to fungal pathogens in rice</article-title>. <source>Planta</source><volume>236</volume>, <fpage>1485</fpage>–<lpage>1498</lpage> (<year>2012</year>).<pub-id pub-id-type="pmid">22798060</pub-id></mixed-citation></ref><ref id="b34"><mixed-citation publication-type="journal">Shimono, <italic>et al.</italic><article-title>Rice WRKY45 plays a crucial role in benzothiadiazole-inducible blast resistance</article-title>. <source>Plant Cell</source><volume>19</volume>, <fpage>2064</fpage>–<lpage>2076</lpage> (<year>2007</year>).<pub-id pub-id-type="pmid">17601827</pub-id></mixed-citation></ref><ref id="b35"><mixed-citation publication-type="journal"><name><surname>Nakayama</surname><given-names>A.</given-names></name><italic>et al.</italic><article-title>Genome-wide identification of <italic>WRKY45</italic>-regulated genes that mediate benzothiadiazole-induced defense responses in rice</article-title>. <source>BMC Plant Biol.</source><volume>13</volume>, <fpage>150</fpage> (<year>2013</year>).<pub-id pub-id-type="pmid">24093634</pub-id></mixed-citation></ref><ref id="b36"><mixed-citation publication-type="journal"><name><surname>Goto</surname><given-names>S.</given-names></name><italic>et al.</italic><article-title>Development of disease-resistant rice by pathogen-responsive expression of WRKY45</article-title>. <source>Plant Biotechnol. J.</source><volume>13</volume>, <fpage>753</fpage>–<lpage>765</lpage> (<year>2015</year>).<pub-id pub-id-type="pmid">25487714</pub-id></mixed-citation></ref><ref id="b37"><mixed-citation publication-type="journal"><name><surname>Wu</surname><given-names>C. J.</given-names></name><italic>et al.</italic><article-title>Inhibition of severe acute respiratory syndrome corona virus replication by niclosamide</article-title>. <source>Antimicrob. Agents Chemother.</source><volume>48</volume>, <fpage>2693</fpage>–<lpage>2696</lpage> (<year>2004</year>).<pub-id pub-id-type="pmid">15215127</pub-id></mixed-citation></ref><ref id="b38"><mixed-citation publication-type="journal"><name><surname>Zhu</surname><given-names>P. J.</given-names></name><italic>et al.</italic><article-title>Quantitative high-throughput screening identifies inhibitors of anthrax-induced cell death</article-title>. <source>Bioorg. Med. Chem.</source><volume>17</volume>, <fpage>5139</fpage>–<lpage>5145</lpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19540764</pub-id></mixed-citation></ref><ref id="b39"><mixed-citation publication-type="journal"><name><surname>Osada</surname><given-names>T.</given-names></name><italic>et al.</italic><article-title>Antihelminth compound niclosamide downregulates Wnt signaling and elicits antitumor responses in tumors with activating APC mutations</article-title>. <source>Cancer Res.</source><volume>71</volume>, <fpage>4172</fpage>–<lpage>4182</lpage> (<year>2011</year>).<pub-id pub-id-type="pmid">21531761</pub-id></mixed-citation></ref><ref id="b40"><mixed-citation publication-type="journal"><name><surname>Balgi</surname><given-names>A. D.</given-names></name><italic>et al.</italic><article-title>Screen for chemical modulators of autophagy reveals novel therapeutic inhibitors of mTORC1 signaling</article-title>. <source>PLoS One</source><volume>4</volume>, <fpage>e7124</fpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19771169</pub-id></mixed-citation></ref><ref id="b41"><mixed-citation publication-type="journal"><name><surname>Chen</surname><given-names>M.</given-names></name><italic>et al.</italic><article-title>The anti-helminthic niclosamide inhibits Wnt/Frizzled1 signaling</article-title>. <source>Biochemistry</source><volume>48</volume>, <fpage>10267</fpage>–<lpage>10274</lpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19772353</pub-id></mixed-citation></ref><ref id="b42"><mixed-citation publication-type="journal"><name><surname>Wang</surname><given-names>A. M.</given-names></name><italic>et al.</italic><article-title>The autonomous notch signal pathway is activated by baicalin and baicalein but is suppressed by niclosamide in K562 cells</article-title>. <source>J. Cell Biochem.</source><volume>106</volume>, <fpage>682</fpage>–<lpage>692</lpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19160421</pub-id></mixed-citation></ref><ref id="b43"><mixed-citation publication-type="journal"><name><surname>Fonseca</surname><given-names>B. D.</given-names></name><italic>et al.</italic><article-title>Structure-activity analysis of niclosamide reveals potential role for cytoplasmic pH in control of mammalian target of rapamycin complex 1 (mTORC1) signaling</article-title>. <source>J. Biol. Chem.</source><volume>287</volume>, <fpage>17530</fpage>–<lpage>17545</lpage> (<year>2012</year>).<pub-id pub-id-type="pmid">22474287</pub-id></mixed-citation></ref><ref id="b44"><mixed-citation publication-type="journal"><name><surname>Weinbach</surname><given-names>E. C.</given-names></name> & <name><surname>Garbus</surname><given-names>J.</given-names></name><article-title>Mechanism of action of reagents that uncouple oxidative phosphorylation</article-title>. <source>Nature</source><volume>221</volume>, <fpage>1016</fpage>–<lpage>1018</lpage> (<year>1969</year>).<pub-id pub-id-type="pmid">4180173</pub-id></mixed-citation></ref><ref id="b45"><mixed-citation publication-type="journal"><name><surname>Prathuangwong</surname><given-names>S.</given-names></name> & <name><surname>Amnuaykit</surname><given-names>K.</given-names></name><article-title>Studies on tolerance and rate reducing bacterial pustule of soybean cultivars/lines</article-title>. <source>Kasetsart J</source>. <volume>21</volume>, <fpage>408</fpage>–<lpage>420</lpage> (<year>1989</year>).</mixed-citation></ref><ref id="b46"><mixed-citation publication-type="journal"><name><surname>Mitsuhara</surname><given-names>I.</given-names></name><italic>et al.</italic><article-title>Characteristic expression of twelve rice PR1 family genes in response to pathogen infection, wounding, and defense-related signal compounds (121/180)</article-title>. <source>Mol. Genet. Genomics</source><volume>279</volume>, <fpage>415</fpage>–<lpage>427</lpage> (<year>2008</year>).<pub-id pub-id-type="pmid">18247056</pub-id></mixed-citation></ref><ref id="b47"><mixed-citation publication-type="journal"><name><surname>Hwang</surname><given-names>S.-H.</given-names></name>, <name><surname>Lee</surname><given-names>I. A.</given-names></name>, <name><surname>Yie</surname><given-names>S. W.</given-names></name> & <name><surname>Hwang</surname><given-names>D.-J.</given-names></name><article-title>Identification of an OsPR10a promoter region responsive to salicylic acid</article-title>. <source>Planta</source><volume>227</volume>, <fpage>1141</fpage>–<lpage>1150</lpage> (<year>2008</year>).<pub-id pub-id-type="pmid">18193274</pub-id></mixed-citation></ref><ref id="b48"><mixed-citation publication-type="journal"><name><surname>Ryu</surname><given-names>H.-S.</given-names></name><italic>et al.</italic><article-title>A comprehensive expression analysis of the WRKY gene superfamily in rice plants during defense response</article-title>. <source>Plant Cell Rep.</source><volume>25</volume>, <fpage>836</fpage>–<lpage>847</lpage> (<year>2006</year>).<pub-id pub-id-type="pmid">16528562</pub-id></mixed-citation></ref><ref id="b49"><mixed-citation publication-type="journal"><name><surname>Liua</surname><given-names>X.</given-names></name>, <name><surname>Baia</surname><given-names>X.</given-names></name>, <name><surname>Wanga</surname><given-names>X.</given-names></name> & <name><surname>Chua</surname><given-names>C.</given-names></name><article-title>OsWRKY71, a rice transcription factor, is involved in rice defense response</article-title>. <source>J. Plant Physiol.</source><volume>164</volume>, <fpage>969</fpage>–<lpage>979</lpage> (<year>2007</year>).<pub-id pub-id-type="pmid">16919842</pub-id></mixed-citation></ref><ref id="b50"><mixed-citation publication-type="journal"><name><surname>Jin</surname><given-names>Y.</given-names></name><italic>et al.</italic><article-title>Antineoplastic mechanisms of niclosamide in acute myelogenous leukemia stem cells: inactivation of the NF-kappa B pathway and generation of reactive oxygen species</article-title>. <source>Cancer Res.</source><volume>70</volume>, <fpage>2516</fpage>–<lpage>2527</lpage> (<year>2010</year>).<pub-id pub-id-type="pmid">20215516</pub-id></mixed-citation></ref><ref id="b51"><mixed-citation publication-type="journal"><name><surname>Sack</surname><given-names>U.</given-names></name><italic>et al.</italic><article-title>Novel effect of antihelminthic niclosamide on S100A4-mediated metastatic progression in colon cancer</article-title>. <source>J. Natl. Cancer Inst.</source><volume>103</volume>, <fpage>1018</fpage>–<lpage>1036</lpage> (<year>2011</year>).<pub-id pub-id-type="pmid">21685359</pub-id></mixed-citation></ref><ref id="b52"><mixed-citation publication-type="journal"><name><surname>Vleesschauwer</surname><given-names>D. D.</given-names></name>, <name><surname>Xu</surname><given-names>J.</given-names></name> & <name><surname>Höfte</surname><given-names>M.</given-names></name><article-title>Making sense of hormone-mediated defense networking: from rice to <italic>Arabidopsis. Front</italic></article-title>. <source>Plant Sci.</source><volume>5</volume>, <fpage>611</fpage> (<year>2014</year>).</mixed-citation></ref><ref id="b53"><mixed-citation publication-type="journal"><name><surname>Tao</surname><given-names>Z.</given-names></name><article-title>A pair of allelic <italic>WRKY</italic> genes play opposite roles in rice-bacteria interactions</article-title>. <source>Plant Physiol.</source><volume>151</volume>, <fpage>936</fpage>–<lpage>948</lpage> (<year>2009</year>).<pub-id pub-id-type="pmid">19700558</pub-id></mixed-citation></ref><ref id="b54"><mixed-citation publication-type="journal"><name><surname>Iwata</surname><given-names>M.</given-names></name><italic>et al.</italic><article-title>Effect of probenazole on the activities related to the resistant reaction in rice plant</article-title>. <source>Ann. Phytopathol. Soc. Jpn</source>. <volume>46</volume>, <fpage>297</fpage>–<lpage>306</lpage> (<year>1980</year>).</mixed-citation></ref><ref id="b55"><mixed-citation publication-type="journal"><name><surname>Friedrich</surname><given-names>L.</given-names></name><italic>et al.</italic><article-title>A benzothiadiazole derivative induces systemic acquired resistance in tobacco</article-title>. <source>Plant J.</source><volume>10</volume>, <fpage>61</fpage>–<lpage>70</lpage> (<year>1996</year>).</mixed-citation></ref><ref id="b56"><mixed-citation publication-type="journal"><name><surname>Görlach</surname><given-names>J.</given-names></name><italic>et al.</italic><article-title>Benzothiadiazole, a novel class of inducers of systemic acquired resistance, activates gene expression and disease resistance in wheat</article-title>. <source>Plant Cell</source><volume>8</volume>, <fpage>629</fpage>–<lpage>643</lpage> (<year>1996</year>).<pub-id pub-id-type="pmid">8624439</pub-id></mixed-citation></ref><ref id="b57"><mixed-citation publication-type="journal"><name><surname>Lawton</surname><given-names>K. A.</given-names></name><italic>et al.</italic><article-title>Benzothiadiazole induces disease resistance in Arabidopsis by activation of the systemic acquired resistance signal transduction pathway</article-title>. <source>Plant J.</source><volume>10</volume>, <fpage>71</fpage>–<lpage>82</lpage> (<year>1996</year>).<pub-id pub-id-type="pmid">8758979</pub-id></mixed-citation></ref><ref id="b58"><mixed-citation publication-type="journal"><name><surname>Francesco</surname><given-names>I.</given-names></name><italic>et al.</italic><article-title>New life for an old drug: the anthelmintic drug niclosamide inhibits <italic>Pseudomonas aeruginosa</italic> quorum sensing</article-title>. <source>Antimicrob. Agents. Ch</source>, <volume>57</volume>, <fpage>996</fpage>–<lpage>1005</lpage> (<year>2013</year>).</mixed-citation></ref><ref id="b59"><mixed-citation publication-type="journal"><name><surname>Ofori-Adjei</surname><given-names>D.</given-names></name>, <name><surname>Dodoo</surname><given-names>A. N. O.</given-names></name>, <name><surname>Appiah-Danquah</surname><given-names>A.</given-names></name> & <name><surname>Couper</surname><given-names>M.</given-names></name><article-title>A review of the safety of niclosamide, pyrantel, triclabendazole and oxamniquine</article-title>. <source>Int. J. Risk Saf. Med</source>. <volume>20</volume>, <fpage>113</fpage>–<lpage>122</lpage> (<year>2008</year>).</mixed-citation></ref><ref id="b60"><mixed-citation publication-type="journal"><name><surname>Park</surname><given-names>B. S.</given-names></name>, <name><surname>Song</surname><given-names>J. T.</given-names></name> & <name><surname>Seo</surname><given-names>H. S.</given-names></name><article-title>Arabidopsis nitrate reductase activity is stimulated by the E3 SUMO ligase AtSIZ1</article-title>. <source>Nat. Commun.</source><volume>2</volume>, <fpage>400</fpage> (<year>2011</year>).<pub-id pub-id-type="pmid">21772271</pub-id></mixed-citation></ref></ref-list><fn-group><fn><p><bold>Author Contributions</bold> S.I.K. and H.S.S. designed the studies. S.I.K. performed experiments. S.I.K., J.T.S., J.Y.J. and H.S.S. interpreted data. S.I.K. and H.S.S. wrote the manuscript. All authors commented on the results and the manuscript.</p></fn></fn-group></back><floats-group><fig id="f1"><label>Figure 1</label><caption><title>Effects of niclosamide on the growth of bacterial and fungal pathogens.</title><p>(<bold>a</bold>) <italic>Xoo</italic> strains PXO99, 10208, and K3a, and a <italic>Xag</italic> strain were grown on PSA medium containing different niclosamide concentrations. (<bold>b</bold>) Growth inhibitory effect of niclosamide on twelve <italic>M. oryzae</italic> strains: 46531, 46532, 46534, 46535, 46536, 46538, 46540, 46541, 46542, 46544, KI215, and PO6-6. (<bold>c</bold>) Effects of parthenolide against strains PXO99 and 10208, which were used as controls. (<bold>d</bold>) Effects of niclosamide on the growth of three different Gram-negative <italic>E. coli</italic> strains: Top10, Rosseta2, and DH10b. (<bold>e</bold>) Minimum inhibitory concentration (MIC) of niclosamide against <italic>Xoo</italic> strains PXO99 and 10208.</p></caption><graphic xlink:href="srep21209-f1"></graphic></fig><fig id="f2"><label>Figure 2</label><caption><title>Niclosamide dosage effects on rice disease responses to the representative <italic>Xoo</italic> strain PXO99.</title><p>(<bold>a</bold>) Leaves were inoculated with PXO99 bacterial suspension using the leaf-clipping method and the plants sprayed with different niclosamide concentrations. Photographs were taken at 16 days post-inoculation. (<bold>b</bold>) Lesion development was examined by measuring lesion length in PXO99-treated plant leaves or niclosamide- and PXO99-treated plant leaves. Standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in lesion lengths (*<italic>p</italic> < 0.01; **<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between untreated and niclosamide-treated leaves. (<bold>c</bold>) Leaves of PXO99-treated plants or niclosamide- and PXO99-treated plants were sampled at the indicated time points. The samples were ground, suspended in sterile water, and plated onto peptone-sucrose agar medium containing cephalexin. The numbers of bacterial colonies were then counted. Standard deviations of the means are indicated by vertical bars. <inline-formula id="d33e1096"><inline-graphic id="d33e1097" xlink:href="srep21209-m7.jpg"></inline-graphic></inline-formula>, PXO99 only; <inline-formula id="d33e1099"><inline-graphic id="d33e1100" xlink:href="srep21209-m1.jpg"></inline-graphic></inline-formula>, PXO99 plus 4 μg/ml niclosamide; <inline-formula id="d33e1102"><inline-graphic id="d33e1103" xlink:href="srep21209-m2.jpg"></inline-graphic></inline-formula>, PXO99 plus 8 μg/ml niclosamide; □, PXO99 plus 20 μg/ml niclosamide; <inline-formula id="d33e1105"><inline-graphic id="d33e1106" xlink:href="srep21209-m3.jpg"></inline-graphic></inline-formula>, PXO99 plus 40 μg/ml niclosamide. Standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in bacterial populations (***<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between untreated and niclosamide-treated leaves at different time points after PXO99 inoculation.</p></caption><graphic xlink:href="srep21209-f2"></graphic></fig><fig id="f3"><label>Figure 3</label><caption><title>Niclosamide effects on rice disease responses to the representative <italic>Xoo</italic> strain PXO99.</title><p>(<bold>a</bold>) Leaves were inoculated with bacterial suspension using the leaf clipping method and sprayed with 8 μg/ml niclosamide. (<bold>b</bold>) Lesion development was examined by measuring lesion length using the leaves of PXO99-treated plants or niclosamide plus PXO99-treated plants. The standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in lesion lengths (***<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between untreated and niclosamide-treated leaves. (<bold>c</bold>) Leaves of PXO99-treated plants or niclosamide- and PXO99-treated plants were sampled at the indicated time points. Samples were ground, suspended in sterile water, and plated onto peptone-sucrose agar medium containing cephalexin. The numbers of bacterial colonies were then counted. Standard deviations of the means are indicated by vertical bars. <inline-formula id="d33e1140"><inline-graphic id="d33e1141" xlink:href="srep21209-m8.jpg"></inline-graphic></inline-formula>, PXO99 only; <inline-formula id="d33e1143"><inline-graphic id="d33e1144" xlink:href="srep21209-m4.jpg"></inline-graphic></inline-formula>, PXO99 plus niclosamide. Standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in bacterial populations (*<italic>p</italic> < 0.01; **<italic>p</italic> < 0.001; ***<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between untreated and niclosamide-treated leaves at different time points after PXO99 inoculation.</p></caption><graphic xlink:href="srep21209-f3"></graphic></fig><fig id="f4"><label>Figure 4</label><caption><title>Systemic effects of niclosamide on rice disease responses to the representative <italic>Xoo</italic> strain PXO99.</title><p>(<bold>a</bold>) Leaves were inoculated with PXO99 bacterial suspension using the leaf-clipping method (left panel). Half of the leaves were covered with polythene bags (middle panel) and the remaining leaves were sprayed with niclosamide (right panel). Photographs were taken at 16 days post-inoculation. (<bold>b</bold>) Lesion development was examined by measuring lesion length in PXO99-treated plant leaves or the local and distal leaves of niclosamide- and PXO99-treated plant leaves. Standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in lesion lengths (***<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between untreated and niclosamide-treated local leaves or between untreated and niclosamide-treated systemic leaves. (<bold>c</bold>) Leaves of PXO99-treated plants or local and distal leaves of niclosamide- and PXO99-treated plants were sampled at the indicated time points. Samples were ground, suspended in sterile water, and plated onto peptone-sucrose agar medium containing cephalexin. The numbers of bacterial colonies were then counted. Standard deviations of the means are indicated by vertical bars. <inline-formula id="d33e1183"><inline-graphic id="d33e1184" xlink:href="srep21209-m9.jpg"></inline-graphic></inline-formula>, PXO99 only; <inline-formula id="d33e1186"><inline-graphic id="d33e1187" xlink:href="srep21209-m5.jpg"></inline-graphic></inline-formula>, PXO99 plus niclosamide (distal); <inline-formula id="d33e1189"><inline-graphic id="d33e1190" xlink:href="srep21209-m6.jpg"></inline-graphic></inline-formula>, PXO99 plus niclosamide (local). Standard deviations of the means are indicated by vertical bars. Asterisks indicate statistically significant differences in bacterial populations (**<italic>p</italic> < 0.001; ***<italic>p</italic> < 0.001; Student’s <italic>t</italic>-test) between niclosamide-treated local and systemic leaves at different time points after PXO99 inoculation.</p></caption><graphic xlink:href="srep21209-f4"></graphic></fig><fig id="f5"><label>Figure 5</label><caption><title>Niclosamide detection in distal leaves.</title><p>Leaves of 80-day-old plants were sprayed with 8 μg/ml niclosamide and compared with untreated distal leaves sampled at the indicated time points. Samples were extracted with methanol, and niclosamide levels in the extracts were detected by HPLC. (<bold>a</bold>) HPLC chromatogram. Inset indicates the chemical structure of niclosamide. (<bold>b</bold>) Niclosamide levels at each time point were expressed in graph form. Asterisks indicate statistically significant differences in niclosamide amounts (***<italic>p</italic> < 0.0001; Student’s t-test) between untreated local leaves and niclosamide-treated systemic leaves.</p></caption><graphic xlink:href="srep21209-f5"></graphic></fig><fig id="f6"><label>Figure 6</label><caption><title>Expression analysis of pathogen-related genes in niclosamide-treated leaves.</title><p>RNA samples were extracted from leaves treated with mock (●), 4 μg/ml niclosamide (○), 8 μg/ml niclosamide (▼), 20 μg/ml niclosamide (Δ), and 40 μg/ml niclosamide (■) at four time points (0, 0.5, 12, and 24 h). <italic>OsPR1</italic>, <italic>OsPR10</italic>, <italic>OsWRKY45</italic>, <italic>OsWRKY62</italic>, <italic>OsWRKY71</italic>, <italic>OsHI-LOX</italic>, and <italic>OsACS2</italic> transcripts were amplified by qRT-PCR using gene-specific primers.</p></caption><graphic xlink:href="srep21209-f6"></graphic></fig><fig id="f7"><label>Figure 7</label><caption><title>Salicylic acid content in niclosamide-treated rice.</title><p>Levels of free SA and glucosyl-SA were evaluated in leaves of rice plants treated with 8 μg/ml niclosamide. Samples were harvested and SA and glucosyl-SA were extracted using a methanol-based method. Free SA (<bold>a</bold>) or total SA (<bold>b</bold>) content combined with free SA and glucosyl-SA were analyzed by HPLC. Results are expressed as the means ± S.D. (<italic>n</italic> = 3).</p></caption><graphic xlink:href="srep21209-f7"></graphic></fig><fig id="f8"><label>Figure 8</label><caption><title>Effect of niclosamide on rice growth and development.</title><p>(<bold>a</bold>) Three-week-old plants were sprayed with 8 μg/ml niclosamide every 4 days. After 70 days, the plants were photographed. (<bold>b</bold>) Photographs of leaves from untreated and niclosamide-treated plants. (<bold>c</bold>–<bold>f</bold>) Height, leaf blade length and width, and leaf blade area of leaves from untreated and niclosamide-treated plants were measured. (<bold>g</bold>) SPAD values were examined in leaves of untreated and niclosamide-treated plants. (<bold>h</bold>–<bold>l</bold>) Granule phenotype, number, and weight were evaluated for mature seeds of untreated and niclosamide-treated plants.</p></caption><graphic xlink:href="srep21209-f8"></graphic></fig></floats-group></pmc></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Wicri/Sante/explor/CovidV2/Data/Pmc/Corpus
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 000060 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Pmc/Corpus/biblio.hfd -nk 000060 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Wicri/Sante
|area= CovidV2
|flux= Pmc
|étape= Corpus
|type= RBID
|clé= PMC:4754756
|texte= Niclosamide inhibits leaf blight caused by Xanthomonas oryzae in rice
}}
Pour générer des pages wiki
HfdIndexSelect -h $EXPLOR_AREA/Data/Pmc/Corpus/RBID.i -Sk "pubmed:26879887" \
| HfdSelect -Kh $EXPLOR_AREA/Data/Pmc/Corpus/biblio.hfd \
| NlmPubMed2Wicri -a CovidV2
| This area was generated with Dilib version V0.6.33. |  |