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WILL AUSTRALIAN ENDEMIC PATHOGENS WEAKEN THE MIGHT OF GIANT RAT’S TAIL (GRT) GRASS? Joseph S. Vitelli1, David G. Holdom1, Roger G. Shivas1, Claire Lock1, Yu Pei Tan1, Kaylene Bransgrove1, Annerose Chamberlain1, Natasha Riding1,James Hosking1 and Brett Cawthray2 1Biosecurity Queensland, DAF, Ecosciences Precinct, Brisbane 2Gladstone Regional Council, Miriam Vale May 2019 NSW BIOTASKFORCE GRT Taxonomy / Identification • Genus Sporobolus contains ~ 200 species in tropical and subtropical areas of the world, including Africa, Asia, Australasia, North and South America • In Australia, 18 species are endemic (native) and further 6 species introduced (naturalised) • Only 4 native species (Sporobolus actinocladus, S. caroli, S. mitchellii and S. virginicus) are regarded as favourable fodder species due to their high protein-content • Three of the native species, S. disjunctus, S. latzii and S. partimpatens are considered rare, and • S. pamelae is listed as endangered in Schedule 2 of the Queensland Nature Conservation Act 1992 Peterson, P.M., Romaschenko, K., Arrieta, Y. H., and Saarela, J.M. (2014). A molecular phylogeny and new subgeneric classification of Sporobolus (Poaceae: Chloridoideae: Sporobolinae). Taxon, 63: 1212–1243 3 Sporobolus distribution in Australia Source: Australia Virtual Herbarium 2019 Clade A. Sporobolus indicus complex (23-26 species) Introduced Natives Others S. africanus (Southern and eastern Africa ) *S. minor (Southern America) S. atrovirens (Mexico) S. molleri (Africa) S. berteroanus, S. natalensis (Africa) S. blakei (Australia) S. myrianthus (Africa) S. creber (Australia) S. pectinellus (Africa) **S. elongatus (Australia) S. pseudairoides, (Southern America) S. farinosus (Guam) S. pyramidalis (Africa/Brazil) S. fertilis (Asia) *S. quadratus (Africa) S. festivus (Africa) S. scabriflorus *S. fourcadii (South Africa) S. sessilis (Australia) S. infirmus (Africa) S. stapfianus (Africa) S. jacquemontii (America) S. tenuissimus (Mexico and South America) S. laxus (Australia) S. trichodes (Mexico) Sporobolus indicus complex distribution 10 Species belonging to Sporobolus indicus complex Source: Australia Virtual Herbarium 2019 Plasticity seen within GRT (S. natalensis) 7 Molecular phylogeny of Australian Sporobolus species Utilising molecular tools to better target weedy Sporobolus classical biological control agents and effectively study the genetic diversity of Sporobolus • Conduct DNA studies against all Sporobolus grasses occurring in Australia to determine their relatedness to native species in Australia and to species in the native range 102 7 Sporobolus natalensis Sporobolus jacquemontii Sporobolus actinocladus Sporobolus mitchelli 9 Endemic pathogen surveys 10 Endemic pathogen surveys 11 12 Sterilisation of lesions Pathogen isolation 1. Surface sterilise visibly infected GRT leaves with ethanol. 2. Excise tissue adjacent to lesions and place sections on an agar plate with antibiotics. 3. Subculture emergent fungi to create a pure culture. 4. Send a sample to Brisbane’s Plant Pathology Herbarium (BRIP) for identification. 13 Which pathogen is causing disease symptoms? Up to 20 foliar pathogens found on an infected plant 14 Promising endemic pathogen Host species BRIP No. Pathogen S. fertilis 66084 a Alternaria arborescens S. natalensis 66616, 68299, 69018 1Colletotrichum spp. Sporobolus sp. 68238/9, 68820 1Colletotrichum spp. S. jacquemontii 66086 a, 66087 a, 66085 a Curvularia ravenelii S. natalensis 66088 a Curvularia ravenelii S. natalensis 69020 Curvularia sp. S. natalensis 66081 a Exserohilum rostratum S. natalensis 65635 a 1Fusarium sp. S. natalensis 66083 a Fusarium proliferatum S. natalensis 68300 Gen. nov. S. natalensis 68298 1Microdochium spp. S. natalensis 65649, 67439a 1,2Microdochium sp. nov. S. natalensis 66617 1,2Neopestalotiopsis sp. nov. S. elongatis 68237 1Neopestalotiopsis sp. S. natalensis Nigrospora spp. S. natalensis 66619 Paraphaeosphaeria michotii S. natalensis 66615 1,2Pestalotiopsis sp. nov. S. natalensis 65632 a, 63688 a, b, c 1Phoma sp. S. natalensis 66618 1Septoria sp. S. natalensis 65638 a 1Stagonospora sp. S. natalensis 65466, 66039, 66324, 66325 Ustilago sporoboli-indici 1Species within this genus are known to be pathogenic grass fungi worldwide 15 Koch’s Postulate methodology Inoculation & re-isolation 1. Harvest spores from agar plate using a Tween 20 solution. 2. Use a hemocytometer to measure spore concentration and adjust to 106 spores per mL of Tween 20. 3. Use a pipetter to apply 10uL of inoculum to each GRT seedling, focusing on the leaf axils (growth points). 4. Place the seedlings in a plastic cage under high humidity (28°C, 60% RH) for five days. 5. After 4 weeks, use the isolation methodology to extract and identify fungi from within the leaf. 16 BRIP 65649 BRIP 65649 Microdochium sp BRIP 65649': A colony on ½PDA after 2 weeks, B colony on ½PDA after 1 week, C conidiomata on ½PDA, D conidial masses, E conidiogenous cells, H conidia. Scale bars: 1 cm (A, B); 200 μm (C); all others – 10 μm. Microdochium dawsoniorum BRIP 65649 Microdochium lycopodinum CBS 125585 98 96 Microdochium phragmitis CBS 285.71 Microdochium fisheri CBS 242.91 S. creber S. natalensis Microdochium nivale CBS 116205 62 Microdochium neoqueenslandicum CBS 108926 37 Microdochium novae-zelandiae CPC 29376 100 Microdochium chrysanthemoides LC5363 82 Microdochium colombiense CBS 624.94 59 63 Microdochium musae CBS 143499 Microdochium seminicola KAS3576 63 Microdochium citrinidiscum CBS 109067 Microdochium tainanense CBS 269.76 81 Microdochium trichocladiopsis CBS 623.77 Microdochium dawsoniorum BRIP 65649 100 Microdochium dawsoniorum BRIP 67439a Idriella lunata CBS 204.56 0.04 BRIP 66615 Pestalotiopsis sp. BRIP 66615’: A colony on ½PDA after 2 weeks, B S. creber S. natalensis colony on ½PDA after 1 week, C-D conidiomata on ½PDA, E-G conidiogenous cells, H conidia. Scale bars: 1 cm (A, B); 200 μm (C); 100 μm (D); all others – 10 μm. 18 . BRIP 66617 S. creber S. natalensis Neopestalotiopsis sp. BRIP 66617: A colony on ½PDA after 2 weeks, B colony on ½PDA after 1 week, C-D conidiomata on ½PDA, E-G conidiogenous cells, H conidia. Scale bars: 1 cm (A, B); 100 μm (C); 200 μm (D); all others – 10 μm. 19 Lesion on Sporobolus natalensis leaf sheath 15 weeks post-inoculation with Neopestalotiopsis sp. BRIP 66617 20 Dead S. natalensis seedling one month after ‘Microdochium sp. BRIP 65649' inoculation (right), 'Microdochium sp. BRIP 65649' growth 10 days after re-isolation (left). Scale bars: 1 cm 21 Dead S. natalensis seedling one month after 'Pestalotiopsis sp. BRIP 66615' inoculation (right), 'Pestalotiopsis sp. BRIP 66615' growth 10 days after re-isolation (left). Scale bars: 1 cm 22 Dead S. natalensis seedling one month after 'Neopestalotiopsis sp. BRIP 66617' inoculation (right), 'Neopestalotiopsis sp. BRIP 66617' growth 10 days after re-isolation (left). Scale bars: 1cm 23 Sporobolus species germinant mortality associated with isolate treatment Lock, C (2018). Investigation of fungal pathogens for the biological control of giant rat’s tail grass (Sporobolus natalensis) in Australia. Honours Research Thesis. Bachelor of Environmental Management The University of Queensland, Brisbane. 24 Phoma sp. (preference for mature plants) Pathogen - black stem fungus from dying GRT grass – Fungus collected from GRT plants at Taunton – Identified genus level as Phoma, New recorded species, Based on ITS sequences – Phoma a ubiquitous (present everywhere), cosmopolitan fungus commonly found in soils – Pathogen causes leaf spot, foliar collapse and wilting of the entire plant – Wilted leaves and blackening shoots 1st observed at plant base, with symptoms progressing upward – Phoma known to overwinter in diseased material – Burning paddocks reduces the presences of Phoma GRT Endemic pathogen survey commenced February 2017 GRT paddock with minimal seed heads was found to contain a leaf smut 26 “Unthrifty” GRT plant with leaf smut 27 Leaf smut around Taunton – characterised with black lines on the leaf blades called sori Sori 28 Leaf smut more prominent on young growth – from heavily grazed GRT plants or slashed plants 29 Burning encouraged new growth Unburnt control Leaf smut re- establishing on GRT plants 8 weeks post fire from a nearby paddock infested with leaf smut 30 Leaf smut (Ustilago sporoboli-indici) Ruptured epidermis, releasing a powdery mass of blackish-brown, spores that are stuck together Germinating spores in agar plate Sporobolus stems (circled) with Leaf smut has been found on 17 formation of teliospores (black) properties across Taunton, Conondale, Gin Gin and Bundaberg. 31 GRT paddock with minimal seed heads GRT seed viability following leaf smut (Ustilago sporoboli-indici) infection (in progress) Immature flower heads infected with leaf smut have minimal-viable seed 33 GRT seed production following leaf smut infection Inflorescence damage Inflorescence Length (cm) Seed production/inflor 4 (severely deformed) 3.3 <1 3 (stem inflorescence) 12.2 12 2 (“healthy”) 56 896 1 (control) 44.5 870 34 Summary • This is no silver bullet – but maybe the glimmer of hope • An integrated control program using herbicides, pasture competition, fertilising, pasture spelling will be required • Effective South African biocontrol agents may be found but in the meantime we have at least 1 leaf smut (Ustilalgo sporoboli-indici) and 40 promising pathogens in Australia that appear damaging • Koch’s Postulate has determined that 3 pathogens are pathogenic against GRT – more KP’s to do followed by pathogenicity testing • Mass producing the agent will hopefully be cheap and easy • Landholders noticing die-back of Sporobolus species are encouraged to contact Biosecurity Queensland to help identify potential pathogens 35 Thank you 36.
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    finalreportp NATURAL RESOURCE MANAGEMENT Project code: NBP.337 Prepared by: Dr Bill Palmer Biosecurity Queensland Department of Primary Industries and Fisheries, Queensland Date published: October 2008 ISBN: 978 1 74191 342 2 PUBLISHED BY Meat & Livestock Australia Limited Locked Bag 991 NORTH SYDNEY NSW 2059 Biological control of weedy sporobolus grasses by two host specific agents Meat & Livestock Australia acknowledges the matching funds provided by the Australian Government to support the research and development detailed in this publication. This publication is published by Meat & Livestock Australia Limited ABN 39 081 678 364 (MLA). Care is taken to ensure the accuracy of the information contained in this publication. However MLA cannot accept responsibility for the accuracy or completeness of the information or opinions contained in the publication. You should make your own enquiries before making decisions concerning your interests. Reproduction in whole or in part of this publication is prohibited without prior written consent of MLA. Biological control of weedy sporobolus grasses Abstract This report describes one of the first attempts at biological control of an invasive weedy grass. Five exotic Sporobolus spp., collectively known as the weedy sporobolus grasses, are serious weeds along the eastern seaboard of Australia. They are of extremely low palatability and cattle can not utilise them, and are also invasive and easily spread to new properties and areas. Biological control investigations commenced in 2000 with surveys of southern Africa, where S. pyramidalis, S. natalensis and S. africanus originate. Some 70 phytophagous insect species and 23 plant pathogens were found but only two organisms were considered potential biocontrol agents; the leaf smut Ustilago sporoboli-indici L.
  • A Molecular Phylogeny of the Subtribe Sporobolinae and a Classification Of

    A Molecular Phylogeny of the Subtribe Sporobolinae and a Classification Of

    A molecular phylogeny of the subtribe Sporobolinae and a classication of the subfamily Chloridoideae (Poaceae) P M. P1, 5, K R1, 2, Y H A3, J M. S4 1Smithsonian Institution, Department of Botany, National Museum of Natu ral History, Washington, DC 20013-7012, U.S.A.; email: peterson@si . edu 2M . G. Kholodny Institute of Botany, National Acad emy of Sciences, Kiev 01601, Ukraine; email: kromaschenko@gmail . com 3Instituto Politécnico Nacional, CIIDIR Unidad Durango- COFAA, Durango, C.P. 34220, Mexico; email: yolah54@gmail . com 4Canadian Museum of Nature, Research and Collections, Ottawa, Ontario K1P 6P4, Canada; email: [email protected] 5Author for correspondence; email: peterson@si . edu Abstract . The classication of the subtribe Sporobolinae containing the following six genera is poorly understood: Calamovilfa (ve species endemic to North Amer i ca), Crypsis (11 species en- demic to Asia and Africa), Psilolemma (one species endemic to Africa), Spartina (17 species cen- tered in North Amer i ca), Sporobolus (186 species distributed worldwide), and Thellungia (one species from Africa and Asia). The goal of this study was to reconstruct the evolutionary history of the Sporobolinae using molecular data with increased species sampling. Most species in this sub- tribe have spikelets with a single oret, one- veined (occasionally three- veined) lemmas, a ciliate membrane or line of hairs for a ligule, and fruits with free pericarps (modied caryopses). A phyloge ne tic analy sis was conducted on 161 species (250 samples), of which 134 species were in the Sporobolinae, using nuclear rITS (ribosomal internal transcribed spacer region) 1 and 2 sequences to infer evolutionary relationships.
  • Agrostology; an Introduction to the Systematics of Grasses James P

    Agrostology; an Introduction to the Systematics of Grasses James P

    Humboldt State University Digital Commons @ Humboldt State University Botanical Studies Open Educational Resources and Data 2005 Agrostology; An Introduction to the Systematics of Grasses James P. Smith Jr Humboldt State University, [email protected] Follow this and additional works at: http://digitalcommons.humboldt.edu/botany_jps Part of the Botany Commons Recommended Citation Smith, James P. Jr, "Agrostology; An Introduction to the Systematics of Grasses" (2005). Botanical Studies. 10. http://digitalcommons.humboldt.edu/botany_jps/10 This Grasses: General is brought to you for free and open access by the Open Educational Resources and Data at Digital Commons @ Humboldt State University. It has been accepted for inclusion in Botanical Studies by an authorized administrator of Digital Commons @ Humboldt State University. For more information, please contact [email protected]. A G R O S T O L O G Y An Introduction to the Systematics of Grasses BY James P. Smith, Jr. Professor Emeritus of Botany Department of Biological Sciences Humboldt State University Arcata, California Thirteenth Edition January 2005 Copyright 2005 by James P. Smith, Jr. Department of Biological Sciences Humboldt State University Arcata, California 95521 [email protected] All Rights Reserved No portion of this text may be reproduced in any form or by any electronic or mechanical means, including information storage and retrieval systems, without written permission. First Edition: 1972 Second Edition: 1973 Third Edition: 1976 Fourth Edition: 1978 Fifth Edition: 1979 Sixth Edition: 1983 Seventh Edition: 1994 Eighth Edition: 1995 Ninth Edition: 1996 Tenth Edition: 1998 Eleventh Edition: 2000 Twelfth Edition: 2002 T A B L E O F C O N T E N T S 1: INTRODUCTION .