Five species of Botryosphaeriaceae identified from Eucalyptus grown in the urban habitats of Pretoria, South Africa Draginja Pavlic-Zupanca,b*, Happy M. Malemeb, Brenda D. Wingfieldc, Michael J. Wingfieldb, Bernard Slippersc aBiosystematics
Programme-Mycology Unit, Plant Protection Research Institute, Agricultural Research Council (ARC-PPRI), Private Bag X134, Pretoria, 0121, South Africa bDepartment of Microbiology and Plant Pathology and cDepartment of Genetics, Centre of Excellence in Tree Health Biotechnology, Forestry and Agricultural Biotechnology Institute (FABI), Faculty of Natural and Agricultural Sciences, University of Pretoria, Pretoria, 0002, South Africa
Email:
[email protected]
Introduction 2
CMW 7772 2 CMW 7054
N. ribis
The Botryosphaeriaceae are opportunistic pathogens of numerous woody plants, including Eucalyptus in a variety of different habitats. These fungi cause canker and die-back diseases on non-native Eucalyptus trees in South African plantations. These disease symptoms were observed on Eucalyptus trees grown in a Pretoria arboretum that includes 20 Eucalyptus species (Fig. 1).
N. parvum
11 87
CMW 10122 2 2 CMW 23792 88 CMW 20736 14 1
9 100
76
CMW 9081 1 CMW 994 52 1 CMW 9071
9 54
4
CMW 12446 10 CMW 12445 85 1 CMW 23791 1 CMW 24460 CMW 24571 2 CMW 15952 16 CMW 15953 93 WAC 12398
N. macroclavatum N. eucalypti
Aim
N. vitifusiforme N. ursorum N. luteum N. australe
To identify Botryosphaeriaceae from 20 Eucalyptus species grown in urban areas in Pretoria, and test their pathogenicity an glasshouse trials.
100
STE U5050 3 281 STE U5252 98 1 CMW 23790 8 CMW 24480 100 17 CMW 10309 2 CMW 9076
4
CMW 1110 CMW 9072 1
N. cryptoaustrale
58
CMW 3386 1 CMW 20738 CMW 23787 1 CMW 23785
85 90
100
3 96 16 100
2 71
CMW 6217 CMW 6229
N. eucalypticola
67
100
CMW 10125 6 1 CMW 11705 98
N. eucalyptorum
4
7
100 64 100
CMW 8000 1 CMW 20728 1 CMW 20739 2 CMW 9075 CMW 20718 CMW 20717 1
B. dothidea
70 100
CMW 20732 CMW 23783 1 Lasiodiplodia theobromae CMW 9074 2 Lasiodiplodia theobromae CMW 10130 5 changes
Fig. 2. One of the 88 most parsimonious trees obtained from the combined sequence datasets of the ITS rDNA and EF-1α loci (tree length = 369 steps, CI = 0.805, RI = 0.953). Bootstrap values based on 1000 bootstrap replicates are shown below the branches and number of steps above the branches. Isolates sequenced in this study are in different colors.
80
Fig. 1. The Pretoria arboretum that includes 20 Eucalyptus species.
h gh
Mean lesion length (mm)
70
Materials and Methods Botryosphaeriaceae were isolated from diseased and asymptomatic twigs and leaves collected from 20 Eucalyptus spp. growing in an arboretum in Pretoria, South Africa, as well as from surrounding wild Eucalyptus trees. They were subsequently identified based on anamorph morphology and DNA sequence data for the ITS rDNA, translation elongation factor 1-α (EF-1α) and β-tubulin region. Thirteen isolates representing five species of the Botryosphaeriaceae that were identified in this study were used in pathogenicity tests in the greenhouse, on two-year-old trees of a Eucalyptus grandis clone (ZG-14).
Results and Discussion Five species were identified, including Botryosphaeria dothidea, Neofusicoccum parvum, N. eucalypti, and the more recently described Neofusicoccum ursorum and N. cryptoaustrale (Fig. 2). The latter three were identified for the first time on Eucalyptus in South Africa. Most of these species were collected from the leaves of 17 different Eucalyptus spp., which did not include E. coniocalyx, E. uiminalis and E. obliqua. Neofusicoccum parvum was most commonly isolated and no host specificity was evident for the species of Botryosphaeriaceae. All isolates inoculated on the Eucalyptus clone ZG-14 produced lesions after six weeks. These were significantly larger than those of the controls, confirming their pathogenicity on this host (Fig. 3).
g
60 ef
50
ef
f def
cde
40
bcd
bc b
30
bcd
bc
20 a
10 0
B. dothidea
N. cryptoaustrale
N. eucalypti
N. ursorum
N. parvum
Fig. 3. Mean lesion lengths (mm) for each isolate of five species of Botryosphaeriaceae after inoculation on a Eucalyptus grandis clone (ZG-14). Control inoculations were done with MEA agar. Bars indicate the 95 % confidence limit for each isolate.
Acknowledgements We would like to thank the National Research Foundation (NRF), the Tree Protection Co-operative Programme (TPCP), Department of Science and technology (DST/NRF Centre of Excellence in Tree Health Biotechnology (CTHB), Department of Water affairs and Forestry, University of Pretoria, and Plant protection Research Institute-Agricultural Research Council (ARC-PPRI), South Africa for their financial support.
References Pavlic D, Slippers B, Coutinho TA, Wingfield MJ, 2007. Botryosphaeriaceae occurring on native
Conclusion
Syzygium cordatum in South Africa and their potential threat to Eucalyptus. Plant Pathology
The results of this study showed that variety of Botryosphaeriaceae infect Eucalyptus in the urban habitats of Pretoria. The current study confirmed that a combination of a few common generalists and some rare, apparently specific, species of the Botryosphaeriaceae infect Eucalyptus at any given site. Future studies should focus on native hosts surrounding plots, such as the one studied here, to better understand host and geographic distribution of Botryosphaeriaceae species.
Pavlic-Zupanc D, Maleme HM, Slippers B, Wingfield MJ. 2013. Neofusicoccum ursorum sp. nov.
56:624–636. and Neofusicoccum cryptoaustrale sp. nov. Fungal Planet description sheets:201–202; in Crous PW, Wingfield MJ, Guarro J, et al. Fungal Planet description sheets: 154–213. Persoonia 31:188– 296. Slippers B, Burgess T, Pavlic D, Ahumada R, Maleme H, Mohali S, Rodas C, Wingfield MJ, 2009. A diverse assemblage of Botryosphaeriaceae infect Eucalyptus in native and non-native environments. Southern Forests 71:101–110.
