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Colletotrichum: complex species or species ... - CBS - KNAW

Colletotrichum: complex species or species ... - CBS - KNAW

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The <strong>Colletotrichum</strong> gloeosp<strong>or</strong>ioides <strong>species</strong> <strong>complex</strong>Table 1. (Continued).Species Culture* Host Country GenBank accession numberITS GAPDH CAL ACT CHS-1 GS SOD2 TUB2C. ti ICMP 5285 C<strong>or</strong>dyline australis New Zealand JX010267 JX009910 JX009650 JX009553 JX009897 JX010124 JX010363 JX010441ICMP 4832* C<strong>or</strong>dyline sp. New Zealand JX010269 JX009952 JX009649 JX009520 JX009898 JX010123 JX010362 JX010442C. tropicale MAFF 239933, ICMP 18672 Litchi chinensis Japan JX010275 JX010020 JX009722 JX009480 JX009826 JX010086 JX010318 JX010396<strong>CBS</strong> 124949*, ICMP 18653 Theobroma cacao Panama JX010264 JX010007 JX009719 JX009489 JX009870 JX010097 JX010329 JX010407<strong>CBS</strong> 124943, ICMP 18651 Annona muricata Panama JX010277 JX010014 JX009720 JX009570 JX009868Xanth<strong>or</strong>rhoea preissii Australia JX010261 JX009927 JX009653 JX009478 JX009823 JX010138 JX010369 JX010448C. xanth<strong>or</strong>rhoeae BRIP 45094*, ICMP 17903,<strong>CBS</strong> 127831IMI 350817a, ICMP 17820 Xanth<strong>or</strong>rhoea sp. Australia JX010260 JX010008 JX009652 JX009479 JX009814Glomerella cingulata “f.sp. camelliae” ICMP 10643 Camellia × williamsii UK JX010224 JX009908 JX009630 JX009540 JX009891 JX010119 JX010358 JX010436ICMP 18542 Camellia sasanqua USA JX010223 JX009994 JX009628 JX009488 JX009857 JX010118 JX010357 JX010429ICMP 10646 Camellia sasanqua USA JX010225 JX009993 JX009629 JX009563 JX009892 JX010117 JX010359 JX010437* = ex-type <strong>or</strong> authentic culture, (*) = ex-type <strong>or</strong> authentic culture of synonymised taxon. Sequences downloaded from GenBank, not generated as part of this project are in bold font. Collection abbreviations are listed in the methods.Several <strong>species</strong>-trees analyses were conducted using BEASTv. 1.7.1 (Drummond et al. 2012). Species-trees combine multi-geneand multiple isolate data to reconstruct the evolutionary hist<strong>or</strong>yof hypothesised <strong>species</strong>, rather than individual isolates. BEASTdoes not use concatenation, but rather co-estimates the individualgene trees embedded inside the summary <strong>species</strong> tree. It alsoestimates the time since each <strong>species</strong> shared a common ancest<strong>or</strong>(divergence times). F<strong>or</strong> these analyses the <strong>species</strong> tree ancestralreconstruction option was selected (Heled & Drummond 2010), thegene data partitioned as f<strong>or</strong> BI and the substitution model f<strong>or</strong> eachgene was selected based on the models selected using jModelTest.The individual isolates were grouped into sets of <strong>species</strong> by setting<strong>species</strong> names as trait values. A strict clock was used f<strong>or</strong> theGAPDH gene (as an all intronic sequence it was assumed to beaccumulating mutations at a steady rate) and the other gene clockrates were estimated relative to GAPDH, using an unc<strong>or</strong>relatedlogn<strong>or</strong>mal relaxed clock. The <strong>species</strong> tree pri<strong>or</strong> used f<strong>or</strong> all geneswas the Yule process, with the ploidy type set to nuclear autosomal.Uninf<strong>or</strong>mative pri<strong>or</strong>s were used f<strong>or</strong> all parameters, and wereallowed to auto optimise.The first <strong>species</strong>-tree analysis was conducted using the 158isolate, five gene dataset, with C. boninense and C. hippeastri asthe outgroups. The MCMC chain was set to 1 × 10 8 generationsf<strong>or</strong> the <strong>species</strong> <strong>complex</strong> tree and samples were taken from theposteri<strong>or</strong> every 1000 generations. The analysis was run twiceindependently. The effective sample size (ESS) and traces ofall parameters and convergence of the two runs was checkedwith Tracer and a summary maximum clade credibility <strong>species</strong>tree was built with TreeAnnotat<strong>or</strong> v. 1.7.1 (Drummond et al.2012) using a 10 % burn-in and a posteri<strong>or</strong> probability limit of0.5, setting the heights of each node in the tree to the meanheight across the entire sample of trees f<strong>or</strong> that clade. Separateanalyses were conducted using all eight genes and the samerestricted set of isolates chosen to represent taxa within theMusae clade and the Kahawae clade as were used f<strong>or</strong> the BIanalyses of the eight gene concatenated analyses outlinedabove. F<strong>or</strong> each of the Musae and Kahawae clade analyses,the MCMC chain was set to 5 × 10 7 generations, but otherwiserun as f<strong>or</strong> the five gene dataset.To illustrate the potential limitations of ITS to discriminate <strong>species</strong>within the C. gloeosp<strong>or</strong>ioides <strong>complex</strong>, an UPGMA tree was builtof all 158 ITS sequences, using the Geneious tree builder tool. AUPGMA tree visually approximates a BLAST search, which is basedon distances (and sequence length) rather than c<strong>or</strong>rected nucleotidesubstitutions of m<strong>or</strong>e sophisticated, model-based analyses.Sequences derived in this study were lodged in GenBank (Table1), the concatenated alignment and trees in TreeBASE (www.treebase.<strong>or</strong>g) study number S12535, and taxonomic novelties inMycoBank (Crous et al. 2004).M<strong>or</strong>phologyDetailed m<strong>or</strong>phological descriptions are provided only f<strong>or</strong> those<strong>species</strong> with no recently published description. Few specimenswere examined from infected host material; the descriptionsprovided are mostly from agar cultures. Cultures were grown onDifco PDA from single conidia, <strong>or</strong> from single hyphal tips f<strong>or</strong> the fewspecimens where no conidia were f<strong>or</strong>med, with culture diametermeasured and appearance described after 10 d growth at 18–20 o Cunder mixed white and UV flu<strong>or</strong>escent tubes, 12 h light/12 h dark.Colour codes follow K<strong>or</strong>nerup & Wanscher (1963).www.studiesinmycology.<strong>or</strong>g123

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