COMMENTARY Experimental evidence of innate immunity: a matter of design, convenience or constraints? T. W. J. Garner Institute of Zoology, Regents Park, London, UK Correspondence T. W. J. Garner, Institute of Zoology, Regents Park, London NW1 4RY, UK. Email: trent.garner@ioz.ac.uk doi:10.1111/j.1469-1795.2007.00147.x ‘It is the worst infectious disease recorded among verte- brates in terms of the numbers of species impacted, and its propensity to drive them to extinction’ (ACAP 2005). This statement is part of the Amphibian Conservation Action Plan, the result of the Amphibian Conservation Summit held in Washington, DC in 2005. Granted a biased group if there ever was one (to my knowledge, all attendees were amphibian biologists), and granted actually providing sta- tistically defensible evidence of extinction rather than in- voking it is a tricky proposition (Solow, 2005), the statement is not as bombastic as an initial reading suggests. The disease in question, chytridiomycosis, has been linked to mass mortality events, population decline, local extirpation and possible extinction on multiple continents. The two leading theories regarding the recent emergence of the disease (novel pathogen hypothesis or endemic pathogen hypothesis: Rachowicz et al., 2005) credit global emergence to human-driven climate change or human-mediated move- ment of infected amphibians. A conservation issue if there ever was one. Doug Woodhams et al. (2007) present another in a series of studies examining how proteins secreted by the skin of amphibians may correlate with responses to the agent of chytridiomycosis, Batrachochytrium dendrobatidis (Bd). Examining innate immune responses directly addresses both of the aforementioned hypotheses; besides, such re- search could also provide information crucial to many of the conservation actions that have been proposed as responses to Bd emergence (e.g. prioritizing Bd-sensitive species for captive breeding programmes). Furthermore, if skin peptide secretions are a true measure of susceptibility to infection, information on species-specific peptide profiles would cer- tainly guide modelling the dynamics of Bd in amphibian communities. Immunity, be it innate or acquired, is adaptive and there- fore expected to vary among lineages (Schmid-Hempel, 2003): genes encoding amphibian antimicrobial peptides exhibit the signal of diversifying selection (Duda, Vanhoye & Nicolas, 2002). Research on other amphibian pathogens has shown that genotype, family and even source population can be used to explain responses to pathogen insult in amphibian hosts (Gantress et al., 2003; Pearman & Garner, 2005) and evidence for variation in immune response to Bd within a species may be inferred from the presence of endemic infection after a presumed Bd-driven decline (Re- tallick, McCallum & Speare, 2004) and variation of viru- lence among isolates of Bd (Berger et al., 2005). Woodhams et al. (2007) detected differences in host survival, histology- based estimates of prevalence and the ability of skin peptides isolated from frogs to inhibit Bd growth, and all three responses correlated, more or less. However, exactly what lineage was tested is questionable, as the basic challenge experiment was carried out using animals from a single clutch per species. Thus, species was irretrievably con- founded with matriline and the conclusion that species in this study vary in their ability to survive Bd must be treated with caution. It is unclear if in vitro Bd inhibition assays were also performed using full siblings, as it is not reported in the paper. Nevertheless, the finding that functional variation in putative innate responses is present opens the door for future research that accounts for lineage, age, condition and environment. Woodhams et al. (2007) also reported variation in the amount of circulating polymorphonuclear leucocytes, or granulocytes, detected between groups of orange-eyed tree frogs dosed with Bd or sham-infected with sterile broth. Again, this experiment suffers design flaws; examples in- clude a lack of independence among replicates (frogs housed in groups) and the use of sterile broth as the negative control rather than filtered media in which Bd was previously grown. Even so, to focus on weaknesses (and I challenge anyone to find an experiment lacking weaknesses) overlooks what might be construed from the experiment. Temperature had no effect on the levels of circulating granulocytes, yet field-based studies show a change in temperature associated with the emergence of chytridiomycosis (Pounds et al., 2006; Bosch et al., 2007). Does this mean that the effects observed Animal Conservation 10 (2007) 418–419 c  2007 The Author. Journal compilation c  2007 The Zoological Society of London 418 Animal Conservation. Print ISSN 1367-9430