PONE-D-21-27570Sea lice (Lepeophtherius salmonis) detection and quantification around aquaculture installations using quantification around aquaculture installations usingPLOS ONE

Dear Dr. Krolicka,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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It was reviewed by two experts in the field, and they have recommended some modifications be made prior to acceptance

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Hope you are keeping safe and well in these difficult times

Thanks

Simon

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Academic Editor

PLOS ONE

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

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Reviewer #1: Krolicka et al. have developed a new assay to specifically detect and quantify sea lice eDNA. The manuscript is well written.

There are few minor points.

Line 43: control aquaculture related diseases

Line 82: Italicize “L. salmonis”

Line 178-185: Can you explain why the L. salmonis naupli or copepodites were directly spiked to the filter rather than spiking them to seawater and conducting filtration afterwords.

Figure 1: Legends are too small

Line 332: “The numbers of MGC…” sentence may be not necessary.

Line 336: as 136

Line 369: production limiting factors

Line 408: to some extent

Line 429-431: The idea of this sentence

Line 467-473: Have you evaluated the temporal variation of L. salmonis eDNA?

Supplementary Figure 2: Pacific

Supplementary Figure 3: ….and 0 copies per µl

Supplementary Figure 5 and 6: Please show the statistical significance

Reviewer #2: The author presents a novel and important validation for an eDNA-based approach to sea lice monitoring. Given the ongoing issue of sea lice infestation on Norwegian aquaculture sites and the elusive nature of unattached infective stages, this may provide a powerful tool for early detection of present/future infestation pressure. Overall, this is an interesting and comprehensive study addressing a question of great ecological and economic importance in Norway. I outline a few specific points below that I hope the author will try to address if relevant to the interpretation of their results.

In the case of samples collected from aquaculture sites, is it possible to discriminate between eDNA from nauplii/copepodite stages and eDNA shed from attached or dead adult/sub-adult lice? Given the potentially large contribution of DNA from these larger lice stages, the presence of high numbers of adult/sub-adult lice may obscure the interpretation of the abundance of infective stages. It may be valuable to discuss briefly the potential contribution of adult/sub-adult DNA from farm samples if the author can estimate from their collected data if they believe this may be relevant to the interpretation of their results.

Lines 46-47: The author outlines their rationale for focusing on L. salmonis, as this is the primary sea lice species impacting Norwegian salmon farms. I am wondering if the author suspects this method could be applied to Caligus spp. Do you think special considerations would be required to apply such a method to Caligus spp. due to their broader host range or differences in lifecycle? Or do you think a similar eDNA-based monitoring method could be developed for members of this sea lice genus using a similar approach to that outlined in this this study?

Lines 63-65: As the author describes, L. salmonis exhibit non-parasitic stages. I was wondering if the author has thoughts on how one might incorporate such information into such an eDNA-based monitoring program and/or if it would be necessary for the effectiveness of this monitoring approach. For example, is it possible that eDNA from these non-parasitic stages could be detected from a nearby farm source but that did not necessarily translate to an elevated infection risk for the focal farm (given the latent period prior to these free-living stages becoming infective)? It sounds like the primary proposed application of this methodology is to assess increases in infestation pressure on fish farms, in which case the presence of non-infective stages would likely be related to the abundance of adult lice at a given site. If the author has considered if/how the presence of these non-infective stages may be incorporated into the application of an eDNA-based monitoring program, it may be worth briefly discussing.

Lines 390-400: The authors discuss ecological explanations for observed differences in depth stratified eDNA detections between sampling seasons. It may be worth discussing potential explanations for this phenomenon related to eDNA dispersal and/or stability. For example, is it possible that depth-dependent eDNA mixing and/or degradation rates were more variable in September than in May, independent of sea lice distributions?

Line 532: "Pacyfic" should be spelled Pacific

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Sea lice (Lepeophtherius salmonis) detection and quantification around aquaculture installations using environmental DNA

PONE-D-21-27570R1

Dear Dr. Krolicka,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Simon Clegg, PhD

Academic Editor

PLOS ONE

Additional Editor Comments:

Many thanks for resubmitting your manuscript to PLOS One

As you have addressed all the comments and the manuscript reads well, I have recommended it for publication

You should hear from the Editorial Office shortly.

It was a pleasure working with you and I wish you the best of luck for your future research

Thanks

Simon