PONE-D-20-17862

Environmental factors and occurrence of horseshoe crabs in the northcentral Gulf of Mexico

PLOS ONE

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Reviewer #1: Partly

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Review for PLOS One: D-20-17862

TITLE: Environmental factors and occurrence of horseshoe crabs in the northcentral Gulf of

Mexico

AUTHORS: Maurice G. Estes, Jr., Ruth H. Carmichael, Xiongwen Chen, Sean Carter

The manuscript describes a study of horseshoe crabs on a set of barrier islands along the northcentral Gulf of Mexico coast toward the western end of the species’ range. As the author’s say, such information and the methods used for collecting such information in low density populations is both valuable in its own right and valuable for what it might say about other low density populations of horseshoe crabs including the at-risk Asian species. Little is known about horseshoe crabs in this region so the data are valuable. Particularly useful elements of this study include an evaluation of the effect of land use/land cover classes on horseshoe crab presence and combining molt, carcass and live animal data to evaluate patterns of presence.

General comments

The results described here are hard to get and certainly it is always important to extract as much as possible from such hard-won data, but some of the analyses and conclusions described in this manuscript seem to be a stretch. First, the islands were visited on 9 occasions in 2012 and 4 occasions in 2013 (based on data in Fig. 4). What is the evidence that these animals are rare? Certainly the population density is not like DE Bay, but how do they compare with other populations in the Gulf of Mexico or other areas such as Maine (at the other end of the species’ range)? Second, while all studies have limitations, this study ignores the fact that the animals were studied from May to September and not during the period when peak numbers of live animals were most likely to be found. Fulford & Haehn 2012 show that on barrier islands in Mississippi Sound including Horn Island (which is one of the islands included in this study) there is a peak of nesting in March and April and Rudloe 1980 (missing from the references) shows clearly that there is a major nesting peak in March and April along the northern Gulf coast of Florida with a second and smaller peak in autumn (Sep-Oct). This means that the authors’ central premise that this is a lower density population than elsewhere in the species’ range is questionable. Also, the authors conclude, “The number of live horseshoe crabs was significantly related to day of the year …” but since they did not study the animals during their peak activity, they are only adding evidence for low activity during summer and a second peak of activity in autumn (Fig. 5). Third, data on size is confounded by not separating males and females for live animals and carcasses (Fig. 6), which means that comparisons with other populations are difficult (data by sex were collected). Fourth, Rudloe 1980 found that water inundation (tide + wind surge) was the most important influence on horseshoe crab numbers in the northeastern Gulf, but the present study does not collect data on actual water levels (which might be available from NOAA). Wind speed and direction are recorded, but not how they influence water height. Fifth, environmental variables such as temperature and salinity are averaged over the study period and used to examine the effects of environmental variables on total observed numbers (“Mean environmental values were computed for each variable for survey years 2012, 2013 and for combined 2012-2013 data for the study area.”) rather than using the data from the date (or week) in which the animals were found. The data on numbers of individuals almost certainly included many zeros on survey dates, which means that a different kind of analysis is needed for data of this type. Sixth, a very interesting addition to the study presented here is the molt transport analysis: 65 tagged molts were placed in the environment at Dauphin Island and 65 at Petit Bois with a reported recapture rate of 2.5% (or about 3 individuals). However, from Fig. 8 (the Fig. numbers do not match the text) it looks as though five tagged molts were recaptured (except that 5 of 130 is 3.9%) so the reader is left wondering about the results of this experiment. It is not clear what the line on Fig. 8 represents. Finally, the strongest results in this study come from the correlations between horseshoe crab abundance and land cover/use categories. Since the environmental variables part of the study revealed little (and may not be analyzed correctly), the authors might consider focusing on these LCLU results. In fact, most of the discussion (lines 518-594) focuses on the parts of this study that are strongest.

Editorial comments

Line 32. The abstract begins with a bit of hyperbole: “This study provides the first regional-scale data…” The study covers a roughly 55 mile stretch along the north-central coast of the Gulf of Mexico with horseshoe crab habitat extending farther to the west (eg. barrier islands of Mississippi) and farther to the east (eg. Escambia County, Florida).

Line 62. “Numbers of horseshoe crabs are reportedly higher to the east along the Florida panhandle…” this would be a good point to reference Rudloe 1980.

Line 77. “… in the northwestern Gulf of Mexico” Except that as you know they are found in microtidal environments.

Line 89. “…where horseshoe crab populations are already sparse and…” the population density in this area is not actually known

Line 105 -114. This paragraph is all about remote sensing. Is this a study using remote sensing?

Line 119 sentence ending, “and most adults thought to migrate from shelf or offshore waters seasonally….” needs a citation.

Line 172. Should be “… discharge in winter and spring than in summer and fall…”

Line 186. Should be “…Surveys were conducted from May to August at biweekly and…”

Lines 189-192. “Thirty-two surveys were completed in 2012 (7 each on Dauphin Island and Fort Morgan Peninsula and 9 each on Petit Bois Island and Horn Island) and 15 surveys were completed in 2013 (4 each on Fort Morgan peninsula, Petit Bois Island and Horn Island, and 3 on Dauphin Island)…” This description does not square with the data given in Fig. 4, which amounts to 9 occasions in 2012 and 4 occasions in 2013. Since not all sites were visited each time, it is not possible to tell on Fig. 4 whether no animals were found or the island was not visited. When exactly did the surveys take place at each site? Fig. 5 presents the same data just for live animals but does not separate the sites. Is effort taken into account in Fig. 5?

Lines 192-194. To continue, the authors state, “…ensuring that a sufficient number of surveys were completed under different environmental conditions to relate horseshoe crab counts to environmental co-variates at each site.” But since many of the surveys are on the same dates, as appears to be the case in Fig. 4 & 5, there are really only 13 different dates or data points to compare with the environmental data. Is this a sufficient sample size? Is this a sufficient sample size to come to conclusions such as horseshoe crab “…presence…[is] mediated by a combination of distance from areas of high freshwater discharge…”

Lines 80-90. Of course a paper can’t cite all relevant publications, but this discussion of the factors influencing horseshoe crab distributions does not reference a number of highly relevant and directly related studies (e.g. Vasquez et al., Cheng et al.)

Line 207. Should be “…Calipers were used for smaller juvenile molts to determine prosomal width…”

Line 247. These are climatic means for Dauphin Island.

Line 252-267. This is not clear. How does this give the height of the ocean floor, relative to what?

Line 289-290. “…to constitute an area large enough that correlations with numbers of live horseshoe crabs, molts or carcasses would not be random.” What is that area? A reference is needed here.

Line 292. Fig. 2. The LCLU classes are a valuable part of this study but this figure is not particularly illuminating. Do all the classes actually occur at the four sites? If some are not included then simplify the multiple colors of the legend. We are told that the horseshoe crabs are on the north sides of the islands but this is not clear from the figure and you can’t see that there is a different LCLU on the north side. The figure legend is not clear. It says “…land cover land use classes resampled from 30 m to 300 m by site for horseshoe crab shoreline habitat analysis with survey data from 2012 and 2013.”

Line 359. Should be “…Islands, and during 2013 99% of all juvenile molts were found on Petit Bois Island…”

Line 362. Table 2. It is not clear what the wind direction and wind speed numbers refer to. They appear to be a mean of the values collected from each island (May-Sep). Is this the number that is used in the analyses (as suggested by the methods, lines 312-313)? If so, then this is a very course way to analyze the data; analyzing the numbers found during each survey (with associated environmental conditions) would seem a more appropriate approach. A southwest wind is said to have an effect on numbers (positive or negative?) but the reader is not told how this relates to the beaches. Which brings up the point that taking means of directions is problematic since 359 is right next to 01. Circular distributions require different stats (not arithmetic mean).

Line 396, 402-404. “… placement, and as of 2019 no additional tagged molts were recovered (Fig 6).” This point is not illustrated with Fig. 6. Maybe Fig. 8?

Line 404. Should be “… counted on beach surveys were correctly associated with the site where they were found.”

Line 407. “Live horseshoe crabs ranged in size from 152 to 280 mm (196 ± 5.01), carcasses ranged in size from 60 to 300 mm (193 ± 2.28 mm),…” Are these adults? Are they males or females? It would be much easier to compare these data with other populations if they were separated into males and females.

Line 415. Are the carcasses adults? A mixture of adults and juveniles? What proportion are adults? Are they male or female?

Line 416. Not sure how these peaks are determined in Fig. 6. Should these be presented as frequencies instead of raw numbers – the legend says frequency but the axis says number?

Line 420. Size data are in Fig. 6 not 7. The adult size data need to be separated by sex. Combining everything does not allow comparisons with other sites.

Line 449 “Fig 8. Daily mean discharge data from the Mobile River into Mobile Bay” these data are given in Fig. 9. Since daily discharge numbers are not related to daily or weekly horseshoe crab numbers, I don’t think this figure is particularly useful. The central point is that the two years differ, which is given in the text with numbers.

Line 464. Should be “The number of live horseshoe crabs was significantly related…”

Line 503. “Accordingly, Petit Bois Island, which is more remote from Mobile Bay…” might be better to say, “Consistent with this view” or some such that is not such a strong conclusion – there are lots of possible reasons for these relationships only some of which were measured in this study. And in fact, the environmental part of this study found few correlations. More comparisons with Fulford & Haehn 2012 and Rudloe 1980 and possibly Brockmann & Johnson 2011 would be useful (size, sex ratios, seasonality).

Line 509. “toward the limit of their distribution in Mississippi.” Should be Louisiana since they are found in Chandeleur Islands.

Line 553. “… has limited reproductive value [3].” Might be better to say limited value in population recruitment (RV is often used in a very different context).

Line 557. “While horseshoe crab size has been found to vary with latitude, the size ranges of all animals measured in this study were consistent with previous studies of horseshoe crabs throughout their range.” Well, adult sizes are consistent with some regions, but how do they compare with areas elsewhere in the Gulf of Mexico or elsewhere in their distribution?

Line 583. “In this case, just two years of data substantially increased our knowledge of horseshoe habitat use where limited or no prior studies had been conducted.” Should be “In this case, just two years of data increased our knowledge of horseshoe crab habitat use…” since Fulford & Haehn was a very fine study.

Line 622. “…anthropogenic pressures [65]. Hence, this study contributes to a growing body of evidence that conservation efforts for horseshoe crabs will benefit from considering local habitat attributes and…” citations to a few of these other studies are needed.

Problems with references (incorrect citations)

8. Smith, D. R., Brockmann, H. J., Beekey, M. A., King, T. L., Millard, M. J. & Zaldivar-Rae, J. 2017. Conservation status of the American horseshoe crab (Limulus polyphemus): a regional assessment. Reviews in Fish Biology and Fisheries, 27, 135-175.

27. Smith, D. R., Brousseau, D. J., Mandt, M. T. & Millard, M. J. 2010. Age and sex specific timing, frequency, and spatial distribution of horseshoe crab spawning in Delaware Bay: Insights from a large-scale radio telemetry array. Current Zoology, 56, 563-574.

50. should be Arthropoda

Missing references

Rudloe, A. 1980. The breeding behavior and patterns of movement of horseshoe crabs, Limulus polyphemus, in the vicinity of breeding beaches in Apalachee Bay, Florida. Estuaries, 3, 177-183.

Cheng, H., Chabot, C. C. & Watson, W. H. 2016. Influence of environmental factors on spawning of the American horseshoe crab (Limulus polyphemus) in the Great Bay estuary, New Hampshire, USA. Estuaries and Coasts, 39, 1142-1153

Vasquez, M. C., Brockmann, H. J. & Julian, D. 2017. Between-population differences in multi-stressor tolerance during embryo development in the American horseshoe crab, Limulus polyphemus. Estuaries and Coasts, 40, 1487-1501.

Reviewer #2: This is a well conducted study...and a well written manuscript. Horseshoe crabs are of such great interest to those who study the ecology of northern Gulf barrier island shorelines. This manuscript provides valuable information which expands current knowledge of barrier island ecology in the northcentral Gulf of Mexico. The research plan was well designed. The field work and data analyses were well executed. This is an interesting study worthy of publication.

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Reviewer #1: No

Reviewer #2: No

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Environmental factors and occurrence of horseshoe crabs in the northcentral Gulf of Mexico

PONE-D-20-17862R1

Dear Dr. Estes, Jr.,

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