Ecological specialisation, especially among parasites infecting a set of host species, is ubiquitous in nature. Host specialisation can be understood as resulting from trade-offs in parasite infectivity, virulence and growth. However, it is not well understood how variation in these trade-offs shapes the overall fitness trade-off a parasite faces when adapting to multiple hosts. For instance, it is not clear whether a strong trade-off in one fitness component may sufficiently constrain the evolution of a generalist parasite despite weak trade-offs in other components. A second mechanism explaining variation in specialisation among species is habitat availability and quality. Rare habitats or habitats that act as ecological sinks will not allow a species to persist and adapt, preventing a generalist phenotype to evolve. Understanding the prevalence of those mechanisms in natural systems is crucial to understand the emergence and maintenance of host specialisation, and biodiversity in general.
In their study "Trait-specific trade-offs prevent niche expansion in two parasites", Lievens et al.  report the results of an evolution experiment involving two parasitic microsporidians, Anostracospora rigaudi and Enterocytospora artemiae, infecting two sympatric species of brine shrimp, Artemia franciscana and Artemia parthenogenetica. The two parasites were originally specialised on their primary host: A. rigaudi on A. parthenogenetica and E. artemiae on A. franciscana, although they encounter both species in the wild but at different rates. After passaging each parasite on each single host and on both hosts alternatively, Lievens et al. asked how host specialisation evolved. They found no change in specialisation at the fitness level in A. rigaudi in either treatment, while E. artemiae became more of a generalist after having been exposed to its secondary host, A. parthenogenetica. The most interesting part of the study is the decomposition of the fitness trade-off into its underlying trade-offs in spore production, infectivity and virulence. Both species remained specialised for spore production on their primary host, interpreted as caused by a strong trade-off between hosts preventing improvements on the secondary host. A. rigaudi evolved reduced virulence on its primary host without changes in the overall fitness trad-off, while E. artemiae evolved higher infectivity on its secondary host making it a more generalist parasite and revealing a weak trade-off for this trait and for fitness. Nevertheless, both parasites retained higher fitness on their primary host because of the lack of an evolutionary response in spore production.
This study made two important points. First, it showed that despite apparent strong trade-off in spore production, a weak trade-off in infectivity allowed E. artemiae to become less specialised. In contrast, A. rigaudi remained specialised, presumably because the strong trade-off in spore production was the overriding factor. The fitness trade-off that results from the superposition of multiple underlying trade-offs is thus difficult to predict, yet crucial to understand potential evolutionary outcomes. A second insight is related to the ecological context of the evolution of specialisation. The results showed that E. artemiae should be less specialised than observed, which points to a role played by source-sink dynamics on A. parthenogenetica in the wild. The experimental approach of Lievens et al. thus allowed them to nicely disentangle the various sources of constraints on the evolution of host adaptation in the Artemia system.
 Lievens, E.J.P., Michalakis, Y. and Lenormand, T. (2019). Trait-specific trade-offs prevent niche expansion in two parasites. bioRxiv, 621581, ver. 4 peer-reviewed and recommended by PCI Evolutionary Biology. doi: 10.1101/621581
After receiving three reviews and having assessed your work, I think that your manuscript would greatly benefit from some revisions. All reviewers agree that your work is original and presents interesting results on the evolution of host specialization. I particularly like the trait-based approach that you used and find the overall analysis sound and well conducted. Most reviewers, including myself, agree with your main conclusions. I am convinced that your work will reach a broad audience interested in the role of trade-offs in the evolution of specialization.
The reviewers ask for some clarifications in the presentation of the study system and of the results. I join them in asking you to revise your manuscript and to carefully address the issues raised during this first round of reviews. A revised version will be suitable for a recommendation.
The main reviewers' comments can be summarized as follows:
- please comment on a possible role of inter-parasite competition during infection of the hosts, provided that multi-infections do occur?
- clarify what is the main mode of genetic adaptation in your experiment: from de-novo mutations or standing variation? and what it implies for the evolutionary dynamics of the traits during the experiment.
- better discuss alternative explanations for the apparent lack of adaptive evolution of spore production; is it a strong trade-off or a lack of genetic variation in that trait? can we interpret this lack of evolution as a negative result?
- enhance the presentation of the ecology of the system and the general literature on the topic.
To those, I am adding a few of my own points:
- please clarify if A. parthenogenetica is an obligate host of A. rigaudi, as suggested by your comment on p4 lines 73-74 ("A. rigaudi has much higher fitness in A. parthenogenetica and cannot persist without this host in the field"); how does this affect A. rigaudi's survival in the lab and the results of your experiment, esp. when A. rigaudi is exposed to the alternate host only?
- similarly, if survival of A. rigaudi depends on presence of A. parthenogenetica, how does the parasite persist in the winter when A. p. is absent? (as per page 14 line 364: "A. parthenogenetica are only present from late spring to fall").
- furthermore, it seems that the two parasites don't have the same ecology, and may not co-occur in nature (p14 line 369); all these details appear late in the manuscript. Please provide a clearer and more complete picture of the ecology of the two species earlier in the manuscript, in the Methods section preferably.
- what is the role of phenotypic plasticity in your experiment? can you discuss your results in terms of evolution of norms of reaction instead of only mean trait values? How plastic are the traits you measured? your results clearly present cases of evolution of reaction norms, with maybe mostly changes in intercepts (mean) than slopes (beside maybe virulence). Please comment.
I would be grateful if you could comment on the points raised here and by the reviewers, providing arguments when you don't agree. Please provide a point-by-point answer with clear reference to the main text to speed up the assessment of your revised version. Thanks!