1. Ten species in one: DNA barcoding reveals cryptic species in the neotropical skipper butterfly Astraptes Fulgerator Paul Hebert, Erin Penton, John Burns, Daniel Janzen & Winnie Hallwachs Proceedings of the National Academy of Sciences 2004 Content: Jan Critique: Emily In

2. The Big Picture The scientific method is the process by which scientists, collectively and over time, endeavor to construct an accurate (that is, reliable, consistent and non-arbitrary) representation of the world. -Frank Wolfs 1. Observation and description of a phenomenon. 2. Formulation of an hypothesis to explain the phenomenon. 3. Use of the hypothesis to predict the existence of other phenomena. 4. Performance of experimental tests of the predictions by several independent experimenters and properly performed experiments.

3. Thinking Critically… No species definition! Instead, refers to “sequence threshold (3%) typically encountered between congeneric species pairs” Typically?

4. Thinking Critically… However, imply the use of the biological species concept  Reproductive isolation Genetic distinctiveness Caterpillar colour patterns Food plant usage Differences in adult morphology

5. Genetic Distinctiveness Mitochondrial DNA is solely maternally inherited!  Females are also the “primary agents of food plant selection”  mtDNA does not account for male gene flow What are the males up to? Hey ladies

6. Caterpillar Colour Patterns “on average, FABOV’s ventral orange is slightly paler than that of SENNOV” Seems rather subjective…

7. Food Plant Usage TRIGO and CELT are the only two ‘species’ that do not share their food plants with others! SENNOV and FABOV share 5 food plants LOHAMP and LONCHO can live on each others’ primary food plants BYTTNER also has own food plant, but only four of them were found!

8. Differences in Adult Morphology “close study of adults, sorted by their caterpillar food plant, showed subtle differences in colour, pattern, size, and wing shape”

9. Thinking Critically… Have Hebert et al. made a good, scientific case for reproductive isolation among their proposed species?

10. Thinking critically… Why the neighbour joining tree? “has the advantage of speed” BUT… taxonomy uses character-based, not distance- based trees Makes union of taxonomy and barcoding difficult Lacks objective criteria to delineate species

11. Thinking Critically… So, Hebert gathered enough barcodes to establish the set of objective criteria to delineate species… “sequence threshold (3%) typically encountered between congeneric species pairs”

12. Thinking critically… … and then didn’t use them. Less than 0.5% sequence divergence “Only two taxa (CELT and TRIGO) have minimum sequence divergences (3.4% and 5.4%)”

13. Intraspecific vs. Interspecific The minimum 3% sequence divergence threshold was based on: Interspecific difference = at least 10x intraspecific variation

14. Thinking critically… Even though 8 out of 10 of the proposed species failed to satisfy Hebert’s own species threshold… …he wrote a paper proposing all 10 as species.

15. Why is my critique important?

16. Science is supposed to be objective DNA barcoding has the potential to be an incredibly useful tool By not being scientifically rigorous, Hebert et al. weaken their own arguments