DOI: 10.5061/dryad.ncjsxkt33
title: Do risk-prone behaviours compromise reproduction and increase vulnerability of fish aggregations exposed to fishing?

description: Human disturbances can prompt natural antipredator behaviours in animals,
 affecting how energy is traded-off between immediate survival and
 reproduction. In our study of male squaretail groupers (Plectropomus
 areolatus) in India's Lakshadweep archipelago, we investigated the
 impact of fishing pressure on anti-predatory responses and reproductive
 behaviours by comparing a fished and unfished spawning aggregation site
 and tracking responses over time at the fished site. Using observational
 sampling and predator exposure experiments, we analysed fear responses
 (flight initiation distance, return time), as well as time spent in
 vigilance, courtship, and territorial defence. Unpaired males at fished
 sites were twice as likely to flee from simulated predators and took
 longer to return to mating territories. Contrastingly, males in the
 presence of females at both sites took greater risks during courtship,
 fleeing later than unpaired males, but paired males at the unfished site
 returned earlier. Our findings suggest that high fishing pressure reduces
 reproductive opportunities by increasing vigilance and compromising
 territorial defence, potentially affecting mate selection cues. Altered
 behavioural trade-offs may mitigate short-term capture risk but endanger
 long-term population survival through altered reproductive investment.
 Human extractive practices targeting reproductive aggregations can have
 disruptive effects beyond direct removal, influencing behaviours crucial
 for population survival.
description: Refer to paper and Supplementary Materials (ESM)
 Methods 2.1.Field sites
 Data were collected from a ‘fished’ squaretail grouper spawning
 aggregation site (Bitra) during the new moon phase in January and from an
 ‘unfished’ site (Site 2) in February 2023 and 2024, in the Lakshadweep
 archipelago (details in electronic supplementary material 1, site
 characteristics). Historical data wasere also used from the fished site
 from January 2013 and 2014 (i.e., when it was in an ‘’unfished’ state,
 [20]). Sampling, site and aggregation characteristics are summariszed in
 table 1. Two prominent modes of active in-water reef fishing were observed
 at the fished site: hook-and-line and spearfishing, both involveing
 fishers free-diving in the water (RK, personal observation, details in
 ESM, electronic supplementary material 1). Anti-predator behavioural
 responses have been demonstrated in response to both fishing scenarios
 [16,17,21]. 2.2.Squaretail grouper spawning
 aggregations Squaretail groupers aggregate at both
 study sites for 5–6 days around the new moon from December to March [20].
 Male groupers arrive 2–3 days earlier, establishing temporary territories
 (5–15 m<sup>2</sup>) that are vigorously defended from
 neighbours. Females typically join a day before the new moon and stay for
 2–3 days after. The mating system is described as a ‘'lek’'
 [20]. Given the males’' stationary behaviour and distinct body
 markings facilitating easy identification, our study focussed on analysing
 male reproductive behaviours. 2.3.Population
 surveys The total aggregation area, including the
 ‘'core’' lekking zone, was marked during snorkel surveys (refer
 to [20] and electronic supplementary material 1, for methods). Groupers
 were surveyed within this core area by two observers (RK, IMK) using
 underwater visual sampling on SCUBA along permanent 50 mx× 10 m belt
 transects. The number of transects varied based on the core area size
 (<em>n</em> = 5 fished site, <em>n</em> = 7
 unfished site). Abundance estimates per transect were used to calculate
 average site-level population density (grouper.500
 m<sup>−2</sup>). Observers also recorded fish lengths (5 -cm
 bins) and sex <em>in situ</em><em>in situ</em>
 [20]. Habitat characteristics such as structural complexity and percentage
 of live coral cover were assessed along three 50 -metre transects at the
 sites (details in electronic supplementary material 2).
 2.4.Behavioural observations Males were
 ‘paired’ with a female, if females were observed within male territories
 during focal follows and if displaying ‘'mate-guarding’'
 behaviours, such as maintaining close contact with females or hovering
 nearby (electronic supplementary material 2: table S1).
 (i) Reactive response: Reactive responses
 were measured at the fished and unfished sites during surveys done in
 2023–2024. To determine the reactive fear response in territorial male
 groupers, we used ‘Flight Initiation Distance (FID)’, i.e., the distance
 at which the focal animal moves to avoid an approaching human observer
 [22]. FID is commonly used as an indicator to test for fish wariness in
 response to fishing [16]. We simulated predation risk by horizontally
 approaching, opportunistically selected territorial males
 (<em>n</em> = 72) on SCUBA at a steady pace from a
 standardiszed distance of five5 metres [23]. The approaching observer
 dropped a marker when the male began to move away and the linear distance
 between the grouper position (prior to movement), and the marker was
 measured by a second observer with a tape measure. Time away from
 territory is known to positively influence territory intrusion in spawning
 territorial damselfish [24]. Therefore, we retreated five5 metres from the
 male territory, immediately after the simulated disturbance to record how
 quickly an individual returns to its territory. In a pilot assessment of
 20 males, over 70% of males returned within a minute, suggesting that
 immediate responses were critical for territory holders. We therefore used
 120 secondsec as a cut-off time in our extended dataset.
 (ii) Proactive response: Focal video
 sampling was used to estimate timed-activity budgets, comparing behaviour
 among the fished and unfished sites in 2023–2024, and within the fished
 site in 2013–2014 &and 2023–2024. Observations were conducted on
 108 territorial males over three days at each site, encompassing a day
 before, during, and after the new moon. Male behaviours were categoriszed
 into three states: ‘'courtship’' (male-–female interactions),
 ‘'territorial aggression’' (male-–male interactions), and
 ‘'vigilance’' (electronic supplementary material 2: table S1).
 The proportion of time spent in each activity was calculated by dividing
 the total time in different states (seconds) by the total video length
 (seconds). To determine whether fishing rather than
 site-level population and environmental factors drove time activity
 budgets, we collected information on focal male body size (cm) and the
 number of males and females in an area of 25 m<sup>2</sup>
 (i.e., competitor and mate density) from the centre of focal male
 territories. Habitat condition (live coral cover) and habitat structure
 (structural complexity) were ranked on a scale of 0–5 for each territory
 (electronic supplementary material 2,: table S2), owing to their influence
 on anti-predator behaviours like shelter-seeking [25,26] and predator
 avoidance [27]. Context data were not available for the historical dataset
 (2013-–2014), which was excluded from the analysis of drivers.
 Statistical analysis (i) Reactive
 response: We used bootstrap resampling
 (<em>R</em> = 1000) to generate 95% confidence intervals to
 compare mean Flight Initiation Distance (FID) and territory return times
 between paired and unpaired males at fished and unfished sites.
 Non-overlapping confidence intervals indicated significant differences
 [28]. Proactive response: We also employed bootstrap resampling to compare
 activity budgets between paired and unpaired males at fished and unfished
 sites and within the fished site (2013–2014 pre-fishing and 2023–2024
 fishing scenario). Ternary plots illustrated trade-offs between courtship,
 aggression, and vigilance. (ii) Drivers of activity
 budgets: Bayesian Zero-and-One Inflated Dirichlet
 (zoid) regressions [29] were used to model the influence of fishing
 status, habitat condition, male size, and social context on time activity
 budgets. The analysis involved 70 individuals surveyed in 2023–2024. All
 analysis was conducted in R (electronic supplementary material 3,: Table
 4table S4).
description: # **Do risk-prone behaviours compromise reproduction and increase
 vulnerability of fish aggregations exposed to fishing?**
 [https://doi.org/10.5061/dryad.ncjsxkt33](https://doi.org/10.5061/dryad.ncjsxkt33) # Description of the data and file structure The dataset consists of 6 data files: **FID.csv**: This dataset contains estimates of reactive anti-predatory responses in male squaretail groupers based on experimental manipulation of predation risk (i.e, predator simulation). Estimates of flight initiation distance (FID) and return times of 117 males from the fished and unfished aggregation site. Also includes Information on courtship status of male (paired, unpaired), body size. File used to generate Figure 1. *Metadata description:* *Date (DD/MM/YYYY): Date the focal observations were made.* *Island: Name of the Island of the squaretail grouper aggregation site.* *ID (numeric): Unique Identification number given to a focal individual.* *size (cm) : Total body size (cm) of the focal individual (tip of the snout to edge of caudal fin). Empty cells indicate data not available.* *sex (male/female): Whether male or female. Empty cells indicate data not available.* *habitat (slope/shelf): Whether focal territory was located on the slope or the shelf of the aggregation site. Empty cells indicate data not available* *FID (metres): Flight Initiation Distance (meters). The distance between the observer and focal individual, when it attempts to move away from the observer. Empty cells indicate data not available return_time (seconds): The time taken (seconds) for the disturbed focal individual to return to it's territory after the observer moves away. Empty cells indicate data not available mates (number): Number of squaretail groupers in a 25meter squared area (5 meter radius) around the focal individual's territory. Empty cells indicate data not available. mates*_cat(no/yes): Whether mates were present or not in the immediate vicinity of focal individual. *Empty cells indicate data not applicable.* *female_size (cm): If mates were females, what was the total body size (cm) of females. Empty cells indicate data not applicable.* *male_size (cm): If mates were males, what was the total body size (cm) of males. Empty cells indicate data not applicable.* **timebudgets.csv**: This dataset contains proactive anti-predatory responses, i.e time activity budgets (the proportion of time in vigilance, courtship and aggression behaviours) from 71 focal males in the fished and unfished sites (sampled in 2023/24) and 31 individuals sampled at the fished site in 2013/14 (i.e in a pre-fishing condition). File used to generate Figure 2. *Metadata description:* *island: Name of island of aggregation site, fished (Bitra), unfished (Site_2)* *date (DD/MM/YYYY): Date of focal observation* *video.id (xyz.MVI): File name of associated focal video* *id (1,2,3...): Unique Identification number given to a focal individual.* *time.start (mm:ss): Start time (minutes) of a behavioural observations.* *time.end (mm:ss): End time (minutes) of a behavioural observations.* *duration (seconds): Total duration of a behavioural observation.* *state: Broad behavioural state of the focal individual (vigilance, courtship or aggression, defined in Electronic supplementary material 2, table 1).* *substate: Specific behaviours associated with the behavioural state (described in Electronic supplementary material 2, table 1).* *substate2: additional details associated with substate* *colour: body colour of focal individual (brown mottled, striped, dark spotted, pale spotted, two tone).* *event: instantaneous response of the focal individual to an interaction with a competitor or mate. Empty cells indicate n/a not applicable because event was not observed* *no.interactors (#): number of individuals the focal individual interacted with. Empty cells indicate n/a data not applicable.* *interactor.size (cm): Total body size (top of snout to edge of caudal fin) of interacting individuals (whether larger, smaller or of equal size to focal individual). Empty cells indicate n/a data not applicable.* *outcome: outcome of interactions for the focal individual. Empty cells indicate n/a data not applicable.* *habitat: dominant substrate of where interaction took place. Empty cells indicate n/a data not applicable.* *notes: additional descriptive observations of the interactions. Empty cells indicate n/a data not applicable.* **focal_context.csv**: This dataset is linked to timebudgets and is combined into a single dataset in the code manuscript_figures.RMD. Contains contextual information of focal individuals at the time of sampling, in terms of body size, individual social context (number of mates and competitors in immediate vicinity), habitat etc. File used in GLM analysis of drivers of behaviours. *Metadata description:* *date:(DD/MM/YYYY): Date of focal observation.* *day (Peak, Peak-1,Peak+1...): Number of days before and after the Peak spawning day (i.e, New Moon night).* *island: Name of island of aggregation site, fished (Bitra), unfished (Site_2).* *video_id (name): name of associated video file.* *video.length:(mm:ss): Total length (minutes: seconds) of the video.* *focal_id (1,2,3...): Unique identification number of focal individual.* *sex (M/F,U): Whether focal individual is male , female or unknown. Empty cells indicate data is not available.* *size (cm): Total body size (cm) of the focal individual (tip of the snout to edge of caudal fin). Empty cells indicate data is not available.* *habitat: Habitat type of focal individual (slope, shelf, sand). Empty cells indicate data is not available.* *live_coral (score: 0-5): Live coral cover visually scored between 0-5, where 1 is bare platform and 5 is 100% coral cover in a 1x1 meter squared digital photoquadrat (see electronic supplementary material 2, table2 for details).Empty cells indicate data is not available.* *structurescore: 0-5: Structural complexity (vertical coral canopy height) scored between 0-5 with 0 as lowest and 5 as highest (see electronic supplementary material 2, table2 for details).Empty cells indicate data is not available.* *sand (score: 0-5): Percentage cover of sand visually scored between 0-5 with 0=0% to 5=100%. Empty cells indicate data is not available.* *neighbour size (cm): Total body size (cm) of neighbouring individuals. Empty cells indicate data is not applicable.* *neighbour(#): Number of neighbours in the immediate vicinity (5meter radius) of focal individuals. Empty cells indicate data is not available.* *predators (species name, #). Empty cells indicate data is not available.* *notes: additional descriptive observations of the focal individual interactions.* **popcen.csv**: This dataset contains estimates of population census of the squaretail grouper aggregation conducted using 5-7, permanent belt transects (50 x 10m) established at the fished (Bitra) site since 2013 and unfished (Site_2) site since 2021,respectively. Observations are conducted over 3-5 days days during the new moon lunar phase in January, February and March each year. Dataset used to generate Table 1 and figures in electronic supplementary material 1 and 2. *Metadata description:* *Date (DD/MM/YYYY): Date the focal observations were made.* *Year (YYYY): Year of annual survey.* *Island: Name of the Island of the squaretail grouper aggregation site (Bitra, Site_2).* *Month: Month of survey (Jan, Feb, March).* *Transect.no: (1-7) ID of 50 x 10m belt transect.* *sex: Whether grouper individuals were M (male), F (female) or U(unknown). Empty cells indicate data is not available.* *size (cm): Total body size (cm) of the male (tip of the snout to edge of caudal fin). Empty cells indicate data is not available.* *Depth (metres): Average depth of the belt transect.* *Zone: Broad reef zone of the belt transect (crest, slope, shelf, sand).* *lunar.day: days since peak spawning (i.e, since the new moon day). Empty cells indicate data is not available.* *tot.abun (#): total number of individuals of a particular size and sex.* **BS_livecoral.csv**: Live coral cover visually estimated from photographs of 1m.squared quadrats (n=5) in three belt transects (50 x 10m) at the fished and unfished site. Data used in Table 1. Empty cells indicate data not available. *Metadata description:* *Date: (DD/MM/YYYY): Date of focal observation.* *island: Name of island of aggregation site, fished (Bitra), unfished (Site_2).* *transect (1-3): ID of 50 x 10m belt transect.* *Quadrat (1-5): ID of 1x1meter quadrat placed every 5m along each transect.* *percentage_lc (%): Percentage cover of live coral in the quadrat.* *percentage_sand (%): Percentage cover of live coral in the quadrat.* **BS_structural complexity.csv**: Vertical canopy height (meters) estimated in-situ, n=10 measurements on three belt transects (50 x 10m) at the fished and unfished site. Data used in Table 1. Empty cells indicate data not available. *Metadata description:* *date:(DD/MM/YYYY): Date of focal observation.* *island: Name of island of aggregation site, fished (Bitra), unfished (Site_2).* *transect_no (1-3): ID of 50 x 10m belt transect.* *depth_cm (cm): 10 random vertical height measurements taken within each transect.* # Code/Software manuscript_figrues.Rmd: R code for the analysis and production of Figures in MS. ESM.Rmd: R code for reproducing figures and data used in Electronic Supplementary Material # Notes run manuscript_figures.Rmd before ESM.Rmd as many objects are called in ESM