Title: [Impact of Capture Stress on Caribbean Reef Sharks] Authors: [Sumaya Nur] Affiliation: Nottingham Trent University, Nottingham, United Kingdom Corresponding Author: [N0937741@my.ntu.ac.uk]
Abstract
There are plenty of physiological indicators of stress in sharks that have been studied extensively in various studies. Blotching is a phenomenon found in sharks during capture with very little research found on its effects as a physiological indicator of stress. It is categorised as a visible discolouration of the ventral surface associated with stress. Caribbean reef sharks (Carcharhinus perezi) were the focus of this study and we set out to study blotching time and see what other factors affected the duration of blotching. Sharks were captured and a number of variables were recorded. This included weight, length, heart rate, air and water temperature, cortisol, capture depth and blotching time. We found that blotching time was influenced by capture depth, with a significant positive relationship between depth and blotching time duration. A statistical comparison of the blotching time in the first and second capture found a significant decrease in blotching time during the second capture. This suggests an adaption to repeated stress which is consistent with previous studies that have shown Caribbean sharks to be more resilient to capture-induced stress. Air and water temperatures did not correlate strongly, despite the Caribbean’s generally warm climate. Our findings show that blotching time may be influenced by factors outside of those studied in this study. Further research would allow a better understanding of the long-term effects of blotching providing further insights into stress mitigation strategies and capture protocol..
Keywords
Capture Effects Blotching Shark Physiology
Introduction
Caribbean Reef Shark (Carcharhinus perezi) The apex predator of the reef ecosystem is the Caribbean reef shark (Carcharhinus perezi). Reaching lengths of up to 3 meters this reef-dwelling shark is found throughout the Western Central Atlantic from North Carolina, the Bahamas, the Gulf of Mexico and the Caribbean Sea to Brazil (Carlson et al. 2021) As an apex predator, the Caribbean reef shark plays a critical role in regulating prey populations in the reef. Despite its importance to the delicate balance of the ecosystem, it is currently considered endangered and on the IUCN red list with a status of near threatened. Despite their reputation, sharks play a vital role in our marine ecosystems, yet they are under threat due to their reputation. Human activity, habitat destruction, climate change and fishing are all factors that are affecting their survival and numbers (Horton et al. 2023). The scientific research conducted on sharks is essential in understanding these magnificent creatures in order to protect their numbers. However, the handling of sharks for research purposes has posed many ethical dilemmas due to the methods used to capture these sharks. The most common method of capture even for research purposes is essentially longline fishing. Sharks are captured using hook and line and either secured to the side of the vessel or hauled onto an observation platform. Once the samples and measurements needed are taken the shark is released. The capture and release of sharks induces physical trauma and physiological stress. In some cases (Brooks et al. 2012) if the stress and trauma are excessive enough then immediate or delayed mortality is possible. The study of shark stress physiology is few and far between although the physiological effects of capture have been studied for several different species, there are many gaps in their research. In this study, we aim to delve into the phenomena of blotching. Blotching is a phenomenon recorded in sharks where the ventral surface of the animal exhibits varying degrees of discolouration. It is clear that capture and the subsequent handling of sharks are significant stressors for sharks, leading to physiological responses that result in blotching. Yet little is known or understood about what factors affect blotching and its duration.
In order to study animal populations, especially in the wild, often involves the capture and subsequent handling of animals. Despite international rules and regulations, as well as ethical standards set by the institutions and scientist conducting the research, there will always be potential risks to animal welfare. (Palmer et al. 2020) concluded that there is always room for improvement in the trapping and study of animals, with more licensing and training needed. In this study, we hope our research will provide a better understanding of the effects of capture on Caribbean sharks and allow better practices to be put into place to improve animal welfare.
The factors assessed in this study are expected to have an effect on the blotching times of the sharks. The depth of capture and repeated exposure to capture are the factors that are most likely to have a significant effect on the duration of blotching times. Additionally, environmental factors such as air and water temperatures are expected to have a relationship and affect each other. The aim of this study is to assess the physiological stress-induced factors that affect sharks during capture, with a focus on the phenomena of blotching.
Methodology
Ethics Statement
In this study, we set out to minimise harm and stress to the animals whilst also ensuring the collection of the data necessary for this study. The Caribbean reef sharks were treated with the utmost regard for their welfare. Sharks were captured using hook and line methods as this is widely recognised as the best method to study sharks. Handling was swift and careful in order to ensure sharks spent the least amount of time out of their habitat. The researchers involved were trained in handling techniques to ensure little discomfort or injury to the sharks. Once sharks were onboard we recorded their data including their physiological states upon capture. This included heart rate, body weight, body length and time taken for blotching. Sharks were released as soon as data was collected to minimise distress. This research adheres to the ethical principles of Nottingham Trent University and international standards for elasmobranch Research. ## Importing Data Rtudio is an integrated development environment for R and a valuable tool for data analysis. All analyses were run in R v.4.1.1 using custom scripts.
The shark data was imported into RStudio and the analyses were conducted using the tidyverse package. We conducted our statistical tests to test the normality and correlations of the data set as well as identifying the means of the data. We visualised the information using scatter graphs.
sharks_1_ %>%ggplot(aes(x=air, ##using ggplot to create a scatter graph showing the correlation between air and watery = water))+geom_point()+theme(axis.text=element_text(size=16),axis.title=element_text(size=16))+geom_smooth(method ="loess", se =FALSE,colour ="red")+labs(title ="The Relationship between Water and Air")
Error: object 'sharks_1_' not found
ggplot(sharksub01, aes(x = sex, y = blotch1, colour = sex, fill = sex)) +geom_boxplot() +labs(title ="Blotching Time by Sex") ##using ggplot to generate a boxplot of blotching time by sex
sharks_1_ %>%ggplot(aes(x=blotch, y = depth))+geom_point()+theme(axis.text=element_text(size=16),axis.title=element_text(size=16))+geom_smooth(method ="loess", se =FALSE,colour ="red")+labs(title ="The Relationship between depth of hooking and blotch time")
Error: object 'sharks_1_' not found
Summary of result
The mean summary of our results is summarised in Figure 2. Heart Rate seems to be the variable with the most range (119-166), with a mean of 141.8 bpm. This large range may indicate differing levels of stress among each shark this may be influenced by other factors, such as the depth at which they were captured. (go into detail about heart rate as a stress indicator in sharks using sources) The sharks also had a wide range of measurements in their body sizes. (weight: 65.1-110.94kg; length: 128.3-291). This significant size difference may have further influences on their stress responses. (more details and sources) Cortisol levels ranged from 50.03 to 112.45 mcg/dL, this is expected and aligns with other studies on the expected stress levels during capture. However, we did have an average capture depth of 50.14 with not much of a range in our min and maximum depths which allows us to look more at other environmental factors as depth remains consistent. (depth and its effects sources) Air and water temperatures remained stable with a mean of 35,54 and 23.02 respectively. The average time for blotching to cover 30% of the ventral surface is approximately 35.13 seconds and the range in this variable was quite narrow (30.78-40.08).
The capture process removes the sharks from their natural environment in the water and exposes them to air. Therefore the relationship between air and water is critical in understanding the environmental factors that sharks face during capture. Our data found that air temperatures ranged from 33 to 38 degrees with a mean of 35.54 (see Figure 2), on the other hand, the range for the water temperature was 20.01 to 25.9 with a mean of 23.02. There is a significant difference in these two variables and this further shows that there is a thermal gradient between air and water, which can have a physiological effect on sharks when they are brought to the surface. The scatter graph showcases a relatively flat LOESS curve, this indicates little to no relationship between air and water temperature. These results are further supported by our Spearman’s rank correlation test.
result <-cor.test(sharks_1_$water, sharks_1_$air,alternative ="two.sided",method ="spearman")
Error: object 'sharks_1_' not found
print(result)
Error: object 'result' not found
Our results showed no significant correlation with a Spearman’s rho of -0.056 and a p-value of 0.2082. These results are slightly unusual as historically the air temperature directly affects the sea surface temperature. The heat in the air heats the water and in regions like the Caribbean where the air is persistently warm the ocean tends to also be warmer (Team 2024). (Osgood et al. 2021) found that several elasmobranch species respond to temperature changes in the air and that this is likely due to metabolic constraints and effects on prey availability. Caribbean reef sharks inhabit warm tropical waters in the Atlantic Ocean around coral reefs in the Caribbean Sea. There are few studies on the direct effect of air and water temperatures on this species. Our results may differ from other studies on the correlation between air and water temperatures due to several factors including the time the study was conducted, the instruments we used and the sample size. ## Effect of multiple capture
Sharks were captured a second time to assess their blotching times upon the second capture. The boxplots in Figure show the distribution of blotching time for male and female reef sharks during the first capture (figure) and the second capture event (figure ). Overall, it seems male reef sharks had a longer blotching time. This is most likely due to the difference in size from Figure 2 we can see the male sharks are larger in length and weight compared to the females this is consistent with research. (Manire et al. 2007) found that cortisol levels in female bonnethead sharks (n = 146, x¯ = 1191 pg/mL) were significantly lower than that of males (P < 0.05).
We then conducted a t-test to compare the blotching times in the first capture (blotch 1) and the second capture (blotch 2) for the 50 individuals. Our results showed a significant difference in blotching times. With a t-value of -17.39, p-value of <2.2e-16 and 49 degrees of freedom. With these p values, we know that the difference we have observed is statistically significant and not due to chance. We found that the mean blotching time during the second capture was approximately 0.93 seconds shorter than the first capture, as shown by the mean difference of -0.9297. The fact that the mean difference is negative shows that the blotching time being shorter in the second capture is consistent amongst all individuals. Our findings show that repeated capture influences the shark’s response and is reflected in our reduced blotching times. However, shorter blotching times may also suggest physiological suppression due to stress. Our results show that repeat capture has a significant effect on sharks, and reduced blotching time may suggest lower stress in subsequent captures however, we don’t know the impact of this on shark health over time. Further research is essential to understand the effects of the phenomena of blotching. (Brooks et al. 2012) found that Caribbean reef sharks do experience acute stress when exposed to longer capture durations but do show recovery over time. This supports our results and explains why blotching time decreases. The Caribbean reef shark ranks as a species more resilient to longline capture. (Mandelman and Skomal 2008) ranked the relative resilience of five species of sharks according to the level of blood acid-base perturbation at the time of longline capture. Their results found the Caribbean reef shark to rank among the most resilient species. This further supports our results and may explain why blotching time decreases. Longer studies with more captures would further explore this. However, this would also raise more ethical concerns as sharks would be subject to capture stress several more times.
Predicting the blotching time
Using the data we collected, we wanted to see if there was a way to predict blotching time perhaps. We generated a scatterplot on r to visualise the relationship between blotching time and hooking depth. ( some data on how depth of hooking affects sharks) The graph shows a very clear positive relationship between blotching time and depth. The loess curve slopes upwards, which indicates that longer blotching times are associated with deeper hooking. To further assess the relationship between blotching time, we completed a linear regression model on r. We tested blotching time against several variables: depth, cortisol levels (meta) and heart rate (bpm).
lm_model <-lm(blotch ~ depth + meta + BPM, data = sharks_1_)
Error in eval(mf, parent.frame()): object 'sharks_1_' not found
summary(lm_model)
Error: object 'lm_model' not found
The linear regression evaluates how the predictor’s depth, cortisol and heart rate explain the variation in blotching time. Our R-squared was 0.5108 therefore, the model explains 51% of the variance a moderate, though there is room for improvement. Just like we predicted with our scatter graph, depth was the best predictor, with a significant positive relationship (p-value< 2e-16). So for every unit increase in depth, blotching time also increases by 0.5046 units. This suggests that the depth at which the sharks are hooked plays a critical role in the time it takes for blotching to cover 30% of the ventral surface. A study by (Morgan and Carlson 2010) found that mortality rates increased in several species of shark with increased time on the hook. The lower the depth the sharks are captured at the longer their time spent on the hook. This may explain why the depth captured had such a significant effect on blotching time On the other hand, the other two predictors, meta and bpm, had insignificant coefficients (p-values of 0.623 and 0.510, respectively). The model also showed a residual standard error of 1.001, which suggests accuracy in predicting blotching. The F statistic (p-value <2.2e-16), however, confirms that, as a whole, the model is statistically significant. In elasmobranchs, cortisol concentrations have been shown to increase following exposure to capture stress (Anderson 2012) However, this may not have a direct effect on blotching as there was no correlation between the two. Further study would help to gain a better understanding of this. In conclusion, the model shows that blotching time can be predicted based on depth. The model needs some improvements in order to make more accurate predictions. Perhaps exploring other predictors might explain more variability in the blotching time as the variables meta and bpm have no significant effect. The model also is limited in generalising predictions beyond our data set.
Conclusion
This study highlights the stress-induced physiological responses of Caribbean reef sharks during capture. Blotching is a critical sign of acute stress and should be further studied to understand its effects on the long-term welfare of Caribbean reef sharks. Blotching was heavily influenced by the depth at which sharks were captured. The deeper hooking were associated with longer blotching times. This suggests that the environmental conditions during capture have a significant effect on shark physiology. In our study cortisol levels and heart rate were not strong predictors for blotching times. We also found that repeated capture had an effect on shark blotching times. There was a significant reduction in blotching times during the second capture. This shows the resilience of the species as supported by other studies (study). Further research is needed however to show the long-term effects on shark welfare. In conclusion, this study has advanced our knowledge of the physiological response to stress in Caribbean reef sharks and has shown that blotching time may be a significant indicator of stress. Further study is needed to explore the additional and long-term effects of blotching as well as examining other factors that could influence blotching. By better understanding stress responses in sharks, we can enhance capture techniques and minimise the affects of capture on animal welfare.
References
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