Seasonal and Nocturnal Activity of Culicoides spp. (Diptera: Ceratopogonidae) Adapted to Different Environments in the Balearic Islands

Round 1

Reviewer 1 Report

This manuscript by Barceló et al. describes efforts to characterize the Culicoides community present throughout the night in an urban and rural site on the Spanish island of Mallorca. The authors used rotating light traps to evaluate nighttime activity of midges from May through November. Overall, the data presented are interesting and do contribute to our knowledge on the overnight flight activity of Culicoides on Mallorca, especially as it pertains to important bluetongue virus vector species. However, I have some concerns about the methodology used and the conclusions that are drawn.

First, as this is framed as a comparison between rural and urban sites in the abstract, it is unfortunate that equal trapping effort was not conducted between the two sites. It’s also unclear whether the trapping that did take place was conducted on the same night (with the same meteorological conditions) or on different nights. It seems misplaced to directly compare these results when the experiment was not designed in such a way. Furthermore, inadequate information is provided on the urban site used, especially as it pertains to variables like light pollution and the impact that may have had on trap efficacy.

My second concern is in the analysis of the data. In general, count data should not be analyzed using linear statistics following a normal distribution. It’s unclear whether any tests of normality were run on the data. The authors should test for normality and either include the outcome in the text or adjust their statistical methods to be more appropriate for the count data being analyzed.

I’m including a few additional notes below:

Introduction:

-Line 44: “…knowledge of ecological and behavioral aspects of this species is still quite limited”. Either this should be changed to “these species” to describe the vector Culicoides species or it should be changed to “this genus” if you’re describing all Culicoides.

-Line 51: I don’t think you can say you determined “annual” activity patterns here since you didn’t trap throughout the year. Your trapping began in May with an already very high abundance of midges present in collections.

Methods:

-Line 61: More information on the “non-farming” property is needed here. In the abstract, this is described as rural vs. urban, but this description of the Rialema property does not adequately convey the characteristics of this urban site. Of particular interest to me is whether there’s significant light pollution present in the urban site compared to the rural site and whether competing light sources could potentially impact the collection outcomes.

-Line 69: Was moon phase data collected during the sample nights? There’s evidence that moon phase can influence trap collections, especially when using only light as a trap bait.

-Line 86: There seems to be text missing here between Nolan et al. and (PCR) assay.

Results:

-Line 123: Were these average temperatures taken from just the nighttime hours during which trapping occurred or is it from the full day?

-A figure or table should be added showing the average temperatures.

Discussion:

-Lines 192-195 are redundant with lines 167-170.

-Lines 211-216: There should also be a mention here of CO2 traps as an intermediary between light baited and animal baited traps.

-Lines 202-205: In some climates, vectors will shift their activity patterns when overnight hours become too cool for host seeking. I think it’s important to specify here that the lack of any daytime data, especially as overnight temperatures cool later in the year, leaves some uncertainty about these activity patterns and potential transmission dynamics.

Figures:

-The figures should be changed to display the actual time periods along the x-axis rather than the collector bottle number. In the methods, you say the collector bottles rotated every 1.5-2 hours, so it’s hard to get a clear picture of the timing of these activity patterns.

Author Response

Please see the attachment

Author Response File: Author Response.pdf

Reviewer 2 Report

The authors present the results of a trapping study aimed at describing the diel and seasonal activity patterns of two main Culicoides vector species (C. imicola and C. obsoletus gp.) in Spain. The authors indicate that these two species have distinct phenologies and activity patterns, which suggest livestock in Mallorca are at risk of BTV for much of the spring and fall. However, I felt the study was somewhat limited in scope, and there are several issues with the paper that need to be addressed.

Editorial Comments

In general, I felt there were several places throughout the manuscript where the phrasing was awkward, or that the word choice was off. I’ve marked them below and tried to provide alternate suggestions when I could.

Line 10: Change “diseases” to “pathogens”

Line 11: Change “affecting to animals” to “affecting animals”

Line 11: Add “virus” after bluetongue

Line 16: Change “characteristic” to “characteristics”

Line 17-20: This sentence is awkward, suggest rephrasing. Maybe make two sentences?

Line 23: Change “avoid” to “control”

Line 28: Change “diseases” to “pathogens”

Line 32: Awkward, suggest rephrasing

Line 36: Change “had” to “have”, add “it” after “when”

Line 39: Add “a” after however

Line 43: Suggest changing “breed” to “rear”

Line 44: Change “species” to “genus”

Line 48: Suggest changing “adapted”

Line 52: Suggest changing “environmental”

Line 104: Suggest changing “in the last” (at the latter?)

Line 123: Remove “registered”

Line 200: Suggest changing “gathering” (peak?)

Line 201: Suggest changing “continental conditions” (mainland?)

Line 203: Remove “us”

General Comments

·      The authors trap at only a single site each at only two locations (one rural and one urban) and so it is difficult to draw any significant conclusions about differences in species diversity or abundance between locations. I don’t think the authors draw any inappropriate conclusions in this regard, but it does limit the scope of the study, especially because they essentially end up with only one site for the analyses because the captures at Rialoma were so low.

·      The authors also do not describe where at each location the traps were set. A high proportion of gravid females at Ca’s Boter and a high proportion of males at Rialema seem to suggest the traps were near a development site, but the authors do not confirm this either through a description of the area or by sampling potential development sites near the traps to confirm the presence of Culicoides larvae.

·      The description of the molecular analysis of the obsoletus complex midges is completely missing from the methods, though there is what appears to be an complete sentence that says “(PCR) assay [18]” in the last paragraph of the methods.

·      Much more detail is needed in the description of the analyses of the data. It’s not clear what hypotheses were tested.

·      No statistics are presented in the results of the paper.

·      Lines 188-190: This statement is not supported by the results of the study and I would suggest removing it

·      Figures 3 and 4 are the exact same

·      The annotation used to describe the differences in collection sizes in table 1 is very confusing. I’m not sure what each month is being compared to.

·      In figures 1 and 2, suggest changing “Collector” to hour or something similar. Even better would be to put hours post sunset or otherwise mark on the figure when sunset and sunrise were

·      Based on figure 1, it appears that maybe the imicola activity peak was missed, and that these midges are active pre-sunset.

Author Response

Please see the attachment

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

This revised manuscript by Barceló et al. describing the activity patterns of Culicoides in urban and rural environments on the Balearic Island of Mallorca has been much improved since the last version. The authors have adequately addressed my initial concerns and comments and I feel the manuscript is nearly ready for publication. I have a few additional minor comments to further improve the manuscript below.

Abstract:

Line 14: adjust “throughout the year” to May-November.

Introduction:

Lines 44-45: I think it would make sense to further elaborate on the current available knowledge of flight patterns, seasonality, and environmental drivers for these species here. You state that they’ve only been partially determined, so provide the reader with some information on what is known at this time that you’re building on with the data you’re collecting in this study.

Methods:

Lines 92-94: I appreciate that you conducted tests for normality when determining whether to use ANOVA here, as you specified in your reviewer response. I think you should include that those tests for normality were used in the text to give confidence in your statistical methods to your readers.

Discussion:

Lines 219-223: I appreciate that this text was added indicating changes in activity as a result of cooler temperatures and reduced daylength. I think the sentence should be updated just slightly to, "some Culicoides adult species enter into diapause or shift their diel activity patterns...". 

Author Response

Please see the attachment

Author Response File: Author Response.pdf