PONE-D-21-15096

First clinical expression of equine insect bite hypersensitivity is associated with co-sensitization to multiple Culicoides allergens

PLOS ONE

Dear Dr. Marti,

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Both reviewers are enthusiastic about your manuscript, however, questions were raised that need to be addressed. In particular, reviewer 1 points out that not all responses to allergens would involve IgE.

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Ulrike Gertrud Munderloh, Ph.D.

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PLOS ONE

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Reviewers' comments:

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Comments to the Author

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Reviewer #1: Partly

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

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: In the manuscript “First clinical expression of equine insect bite hypersensitivity is associated with cosensitization to multiple Culicoides allergens” by Birras et al. the authors explored in a longitudinal study the antigen-specificity of IgE responses to known Culicoides allergens in the serum of horses of islandic breed.

The authors found that almost all horses had shared responses to distinct insect-derived antigens. As responses to most of these antigens were neglectable the season prior to the onset of Insect Bite Hypersensitivity (IBH), the authors conclude that there must be a co-induction to all these antigens at the same time point, during the manifestation of the disease. Alternatively, a dominant antigen could have been the first stimulus that induced an allergic response and then allergic immune responses to all other antigens could have been induced via “epitope spreading”, as it has been shown for many other allergies and auto-immune responses.

Altogether, this is a very nice manuscript, of great interest to the overall scientific community interested in the underlying immunological mechanisms leading to allergies and IBH in specific.

There is just one aspect I would like to see addressed. This is the frequency and strength of allergen-specific non-IgE antibody responses in these animals.

The authors exclusively measured IgE titres in the serum of horses suffering from IBH or not. The half-life of IgE in serum is very short, while IgE remains bound to Ig-ε-R expressing cells for a very long period of time. Thus, serum levels of IgE could easily be misguiding. Furthermore, for many allergens there might also be non-allergic immune reactions, which could induce the expression of allergen-specific IgG antibodies. Such antibodies could potentially outcompete allergen-specific IgE antibodies for binding to the microarray of recombinant antigens, as it has been used in this study. In addition, such antibodies may have a curative effect (e.g. IgG4 antibodies) or may prevent the onset of IBH. For instance, Meulenbroeks et al. (PMID: 25901733) have shown that even horses not affected by IBH are not necessarily immunologically ignorant but have a protective Th1-skewed allergen-specific immune response. Such an alternative explanation could not be ruled out by the presented data. Hence, it appears warranted to complement the current data with total antibody and in particular with allergen-specific IgG antibody titres.

Minor aspect:

Please consistently add “continental” Europe, when referring to Sweden or Switzerland.

Reviewer #2: The paper provides novel information on the development of Equine Insect bite hypersensitivity (IBH) that could be important for the establishment of therapy and prevention of this type of allergy in the future..

The study includes a large number of horses inside every group studied (224 in total) and a control group. A follow-up of three years (enough to obtain good conclusions about the development of the IBH) has been made.

The statistical analyse is well described and is adequate for the data (not normally distribution) and number of animals.

The technique used for the evaluation of the IgE microarray proteins allows the detection of antigenic protein molecules with more precise results than other immunoassay systems used to date and is well described by the authors.

The paper manages to achieve the proposed objectives.

The discussion could be enriched with a broader analysis of the knowledge situation of the IBH so far, but it is understood that the authors have not wanted to enter into comparative aspects because the microarray technique is totally different and much more specific than others used up to now.

I consider that it is suitable to be published.

One error should be corrected on line 40 (same as in the first presentation summary): (<4 years, 4-7 years, > 7 years).

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Reviewer #1: No

Reviewer #2: Yes: Maite Verde, Veterinary Internal Medicine and Dermatology, University of Zaragoza.

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Submitted filename: PONE-D-21-15096 .docx

First clinical expression of equine insect bite hypersensitivity is associated with co-sensitization to multiple Culicoides allergens

PONE-D-21-15096R1

Dear Dr. Marti,

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.

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Kind regards,

Ulrike Gertrud Munderloh, Ph.D.

Academic Editor

PLOS ONE

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