DNA barcoding and evolutionary relationships in Accipiter Brisson,1760 (Aves,

Falconiformes: Accipitridae) with a focus on African and Eurasian representatives

Nomenclature

• A. gentilis: Nl; havik, Eng; gosshawk, Fr; Autour des palombes

• A. nisus: Nl; sperwer, Eng; sparrowhawk, Fr; epervier d’Europe

• Buteo buteo: Nl; buizerd, Eng; Common buzzard Fr; Buse variable

• Buteo albicaudatus; Nl; witstaart buizerd, Eng; white tailed hawk, Fr; Buse à queue blanche

Content• DNA barcoding a short

introduction– General DNA barcoding– Mini barcodes– 10X threshold, best compromise

threshold– BCM and BM criteria– Character based identification

• Coverage– Sample origins– The dataset

• Methods– Character based method (CB)– The 10 X intraspecific distance

threshold (10 X)– The Best compromise threshold

(BCTh)– Best match and best close match

explained

• Results for DNA barcoding– Character based identification– Different thresholds

• Phylogeny reconstruction– 291 bp dataset– 647 bp dartaset

• Discussion• Conclusion

DNA barcoding

• Identification of species with an agreed upon (part of a) gene (COI)

• 5’-3’ part of Cytochrome C Oxidase subunit I

• ~650bp in length

• Polymerase chain reactions and Sanger sequencing

Mini barcodes

• Genetic material from museums is often degraded• Shorter fragments are needed• E.g. sequences of 250-300bp or, in some cases even

shorter, also work for identification purposes

Sonet et al. in press

Sample coverage and origins

• African and European taxa are well represented

• The new world partially represented

• Australia and S-E Asia poorly represented

• Most samples from RMCA/RBINS (n=83/9)

• Supplemented with available genbank sequences (n=28)

sample distribution through time

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<1900 >1900<1930

>1930<1960

>1960<1990

>1990

30-year periodsn

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of

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Series1

The resulting dataset

• 25 European and African Accipiter species with a total of 140 specimens

• sequenced for a 291bp (mini) and 647bp (standard) BC fragment of the COI gene

• three datasets analysed:– dataset A: 25 species, 291bp

– dataset B: 19 species, 647bp

– dataset C: 19 species, 291bp

Character based indentification

– Easy it is about detecting unique differences

– May be unreliable with insufficient sampling

– No threshold whatsoever is used

Average 10 X intraspecific distances and the barcoding gap

• Distances• Arbitrary• Does not always hold

(Tephritidae)• However, works well in

birds

The Best compromise threshold (BCTh)

• Based on the intercept between cumulative intra and interspecific distances

• Based on the dataset under study rather than a predetermined threshold

• Results from one dataset not applicable to another

Lefébure et al. 2006

BM and Best Close match

• Best Match (BM), assignes the species name of its best-matching sequence regardless of how similar the query and reference sequences were. Identification is considered correct when both sequences were from the same species, incorrect if the query species differed from the closest reference species or ambiguous if multiple species were the BM of the query species

• Best close match (BCM) also considers the threshold

BM vs. BCM example

• 4 sequences• 3 species: A. francesiae (n=2) A. badius (n=1) & A. toussenelii (n=1)• BM would assign A. badius and A. toussenelii to the “ambiguous” or

“wrong” (dependent on the threshold) categories• BCM would classify them as impossible when the difference is larger

than the threshold

Results character based identification

Results character based identification

Results BCTh vs. 10 X distance threshold

• Different thresholds

• Partial overlap

• Caused by– A. nisus/rufiventris

– A. virgatus/gularis

– A. cooperii/gundlachi

• At or just below the BC treshold are– A. badius subsp

– A. gentilis subsp

Results BCTh vs. 10 X distance threshold

Results BCTh vs. 10 X distance threshold: Accipiter nisus example

A. rufiventrisA. nisus

Results BCTh vs. 10 X distance threshold: A. Badius and A. brevipes example

Phylogeny reconstruction

• Hypothesis of evolution

• Clades are units of supported taxa (related by direct descent)

• This dataset is only suited for low taxonomic level inferences

• One genegene tree rather than species tree

• Comparison with the current taxonomy based on morphology and morphometrics

Phylogeny based on 291bp

• 11 supported clades– [tachiro]

• A. tachiro subclades

– A. minullus– A. francesiae/ soloensis– [cooperii]– A. badius/ brevipes– A. nisus/ striatus/

erythronemius– A. ovampensis/

madagascariensis– [virgatus] including A.

fasciatus

Phylogeny based on 291bp a closer look

Phylogeny based on 291bp a closer look: Accipiter tachiro example

Phylogeny based on 291bp a closer look: Accipiter badius and A. brevipes example

Phylogeny based on 647bp

• 19 supported clades– [tachiro]

• A. tachiro• A. toussenelii & subsp

– A. soloensis & A. francesiae subsp

– [gentilis]– [cooperii]– [nisus] (without A.

ovampensis/ madagascariensis

– [minullus] (without A. erythropus)

– [virgatus] including A. fasciatus

Phylogeny based on 647bp a closer look

Phylogeny based on 647bp a closer look: The example from Accipiter soloensis and A.

francesiae

Phylogeny based on 647bp a closer look: an example with Accipiter gentilis and A.

melanoleucus

Discussion

• Thresholds– The 10 X threshold would have considered A. brevipes and A. badius,

as well as A. gentilis and A. melanoleucus as belonging to the same species and

• Species concepts– Morphological– Distances based– Phylogenetic

• Support values– Models and interpretation remain issues of discussion

• Sampling– Incomplete or lacking in some cases

Conclusions

• Molecular techniques provide a powerfull method for reviewing taxonomy

• Museum material works!

• The content of the dataset matters

• COI can identify 19 out of the 25 species included

• COI carries a phylogenetic signal suitable for molecular taxonomy on a low taxonomic level

• Parts of genus Accipiter may need revision

Thank you for your attention