Project description:The genus Cercospora includes many important plant pathogens that are commonly associated with leaf spot diseases on a wide range of cultivated and wild plant species. Due to the lack of useful morphological features and high levels of intraspecific variation, host plant association has long been a decisive criterion for species delimitation in Cercospora. Because several taxa have broader host ranges, reliance on host data in Cercospora taxonomy has proven problematic. Recent studies have revealed multi-gene DNA sequence data to be highly informative for species identification in Cercospora, especially when used in a concatenated alignment. In spite of this approach, however, several species complexes remained unresolved as no single gene proved informative enough to act as DNA barcoding locus for the genus. Therefore, the aims of the present study were firstly to improve species delimitation in the genus Cercospora by testing additional genes and primers on a broad set of species, and secondly to find the best DNA barcoding gene(s) for species delimitation. Novel primers were developed for tub2 and rpb2 to supplement previously published primers for these loci. To this end, 145 Cercospora isolates from the Iranian mycobiota together with 25 additional reference isolates preserved in the Westerdijk Fungal Biodiversity Institute were subjected to an eight-gene (ITS, tef1, actA, cmdA, his3, tub2, rpb2 and gapdh) analysis. Results from this study provided new insights into DNA barcoding in Cercospora, and revealed gapdh to be a promising gene for species delimitation when supplemented with cmdA, tef1 and tub2. The robust eight-gene phylogeny revealed several novel clades within the existing Cercospora species complexes, such as C. apii, C. armoraciae, C. beticola, C. cf. flagellaris and Cercospora sp. G. The C. apii s. lat. isolates are distributed over three clades, namely C. apii s. str., C. plantaginis and C. uwebrauniana sp. nov. The C. armoraciae s. lat. isolates are distributed over two clades, C. armoraciae s. str. and C. bizzozeriana. The C. beticola s. lat. isolates are distributed over two clades, namely C. beticola s. str. and C. gamsiana, which is newly described.

Project description:Species delimitation directly impacts on global biosecurity. It is a critical element in the decisions made by national governments in regard to the flow of trade and to the biosecurity measures imposed to protect countries from the threat of invasive species. Here we outline a novel approach to species delimitation, "tip to root", for two highly invasive insect pests, Bemisia tabaci (sweetpotato whitefly) and Lymantria dispar (Asian gypsy moth). Both species are of concern to biosecurity, but illustrate the extremes of phylogenetic resolution that present the most complex delimitation issues for biosecurity; B. tabaci having extremely high intra-specific genetic variability and L. dispar composed of relatively indistinct subspecies. This study tests a series of analytical options to determine their applicability as tools to provide more rigorous species delimitation measures and consequently more defensible species assignments and identification of unknowns for biosecurity. Data from established DNA barcode datasets (COI), which are becoming increasingly considered for adoption in biosecurity, were used here as an example. The analytical approaches included the commonly used Kimura two-parameter (K2P) inter-species distance plus four more stringent measures of taxon distinctiveness, (1) Rosenberg's reciprocal monophyly, (P(AB)),1 (2) Rodrigo's (P(randomly distinct)),2 (3) genealogical sorting index, (gsi),3 and (4) General mixed Yule-coalescent (GMYC).4,5 For both insect datasets, a comparative analysis of the methods revealed that the K2P distance method does not capture the same level of species distinctiveness revealed by the other three measures; in B. tabaci there are more distinct groups than previously identified using the K2P distances and for L. dipsar far less variation is apparent within the predefined subspecies. A consensus for the results from P(AB), P(randomly distinct) and gsi offers greater statistical confidence as to where genetic limits might be drawn. In the species cases here, the results clearly indicate that there is a need for more gene sampling to substantiate either the new cohort of species indicated for B. tabaci or to detect the established subspecies taxonomy of L. dispar. Given the ease of use through the Geneious species delimitation plugins, similar analysis of such multi-gene datasets would be easily accommodated. Overall, the tip to root approach described here is recommended where careful consideration of species delimitation is required to support crucial biosecurity decisions based on accurate species identification.

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Dasyclonium species delimitation

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