The Effects of Pulse Pressure from Seismic Water Gun Technology on ...

Transactions of the American Fisheries Society 142:1335–1346, 2013 ! C American Fisheries Society 2013 ISSN: 0002-8487 print / 1548-8659 online DOI: 10.1080/00028487.2013.802252

ARTICLE

The Effects of Pulse Pressure from Seismic Water Gun Technology on Northern Pike Jackson A. Gross* and Kathryn M. Irvine U.S. Geological Survey, Northern Rocky Mountain Science Center, 2327 University Way, Suite 2, Bozeman, Montana 59715, USA

Siri Wilmoth Wilmoth Statistical Consulting, 308 Scott Street, Post Office Box 621, Gardiner, Montana 59030, USA

Tristany L. Wagner U.S. Geological Survey, Northern Rocky Mountain Science Center, 2327 University Way, Suite 2, Bozeman, Montana 59715, USA

Patrick A. Shields and Jeffrey R. Fox Alaska Department of Fish and Game, Division of Commercial Fisheries, 43961 Kalifornsky Beach Road, Suite B, Soldotna, Alaska 99669, USA

Abstract

We examined the efficacy of sound pressure pulses generated from a water gun for controlling invasive Northern Pike Esox lucius. Pulse pressures from two sizes of water guns were evaluated for their effects on individual fish placed at a predetermined random distance. Fish mortality from a 5,620.8-cm3 water gun (peak pressure source level = 252 dB referenced to 1 µP at 1 m) was assessed every 24 h for 168 h, and damage (intact, hematoma, or rupture) to the gas bladder, kidney, and liver was recorded. The experiment was replicated with a 1,966.4-cm3 water gun (peak pressure source level = 244 dB referenced to 1 µP at 1 m), but fish were euthanized immediately. The peak sound pressure level (SPLpeak ), peak-to-peak sound pressure level (SPLp-p ), and frequency spectrums were recorded, and the cumulative sound exposure level (SELcum ) was subsequently calculated. The SPLpeak , SPLp-p , and SELcum were correlated, and values varied significantly by treatment group for both guns. Mortality increased and organ damage was greater with decreasing distance to the water gun. Mortality (31%) by 168 h was only observed for Northern Pike exhibiting the highest degree of organ damage. Mortality at 72 h and 168 h postexposure was associated with increasing SELcum above 195 dB. The minimum SELcum calculated for gas bladder rupture was 199 dB recorded at 9 m from the 5,620.8-cm3 water gun and 194 dB recorded at 6 m from the 1,966.4-cm3 water gun. Among Northern Pike that were exposed to the large water gun, 100% of fish exposed at 3 and 6 m had ruptured gas bladders, and 86% exposed at 9 m had ruptured gas bladders. Among fish that were exposed to pulse pressures from the smaller water gun, 78% exhibited gas bladder rupture. Results from these initial controlled experiments underscore the potential of water guns as a tool for controlling Northern Pike.

One technology that is being considered as a potential tool for suppressing invasive fish is the seismic water gun (USEPA 2010). In the early 1980s, water guns were developed as an alternative to seismic air guns for seismic exploration (Hutchinson and Detrick 1984). In brief, a water gun operates as a low-energy,

implosive source, producing a cavity formed by the jet of highpressure water expelled from the port end of the gun. Few water guns exist today because water guns were dismissed as a seismic exploration tool. Water guns were less efficient than air guns at producing low-frequency energy and had a greater potential to

*Corresponding author: [email protected] Received August 8, 2012; accepted April 28, 2013

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kill or harm fishes and invertebrates in close proximity to the gun at source levels required for seismic exploration. However, these negative characteristics of water guns may prove useful in the search for suppression tools to combat the threat of aquatic nuisance species. Nonnative aquatic species are altering aquatic and terrestrial communities worldwide (Baxter et al. 2004). For example, generalist predators, such as Northern Pike Esox lucius, have had catastrophic effects on food web structure (Vander Zanden et al. 1999) and ecosystem function (Baxter et al. 2004). Northern Pike are capable of causing potentially irreparable changes in fish community composition (Persson et al. 1996; Bystrom et al. 2007), including the decline and elimination of multiple fish species (e.g., Patankar et al. 2006; Johnson et al. 2008; Muhlfeld et al. 2008). Once established, Northern Pike can be especially damaging to juvenile salmonids, thus threatening the economic and ecological viability of salmonid species (Sepulveda et al. 2013). To date, Northern Pike have had significant negative effects on numerous spawning runs of Coho Salmon Oncorhynchus kisutch, Chinook Salmon O. tshawytscha, and Sockeye Salmon O. nerka from the Susitna River drainage, Alaska (ADFG 2002; Southcentral Alaska Northern Pike Control Committee 2007), and the subsequent expansion of Northern Pike to waterways throughout south-central Alaska has been listed as one of the highest priority threats to wildlife in Alaska (ADFG 2002). The expansion and negative effects of Northern Pike are not specific to Alaska. Nonnative Northern Pike have been implicated in the declines of Atlantic Salmon Salmo salar in Maine (Boucher 2003), trout in Montana (McMahon and Bennett 1996) and California (Vasquez et al. 2012), and endangered fishes in Colorado (Johnson et al. 2008). Northern Pike are currently being removed from systems in northeastern Washington due to growing concern over Northern Pike expansion into the Pacific salmon spawning grounds of the Columbia River basin (D. Osterman, Kalispel Tribe of Indians, personal communication). Primary control methods for reducing invasive Northern Pike in Alaska are piscicides and netting. Numerous other management techniques have been identified and used elsewhere, including water control barriers, dewatering, explosives, electrofishing, stocking of sterilized females, induced winterkill, and biological controls (e.g., Northern Pike predators and parasites). To date, rotenone (a commonly applied piscicide) has been described as the only cost-effective management technique for eradicating Northern Pike from specific water bodies. Although gill nets and piscicides are effective means for reducing the number of Northern Pike, there are several drawbacks and consequences associated with these techniques. For example, nets must be continually monitored to ensure that the bycatch of nontarget fish and aquatic bird species is limited. Piscicides such as rotenone are costly, and their application is labor intensive and can leave traces of chemicals in the surrounding environment. Given these concerns, it is critical to evaluate new

methodologies for preventing and suppressing Northern Pike infestations as part of an integrated management approach to conserve salmon and other game and native fish stocks. Limited dose–response data exist on the effects of pulse pressure from air guns or water guns on fish or the potential effects on other aquatic resources (Popper and Hastings 2009). Therefore, we initiated a set of experiments that were designed to evaluate the feasibility of seismic water gun technology for use as a management tool to suppress Northern Pike. Specific objectives were to (1) assess mortality of Northern Pike as a function of received sound pressure measurements for two consecutive pulses from a 5,620.8-cm3 water gun, with monitoring of mortality over a 168-h posttreatment period; and (2) assess injury thresholds in Northern Pike from exposure to 5,620.8-cm3 and 1,966.4-cm3 water guns as a function of sound pressure measurements. METHODS Study site.—Derk’s Lake (60◦ 19# 46.5594##N, 150◦ 34# 46. 308##W), a mesotrophic lake (surface area = 13.3 ha) near Soldotna, Alaska, was chosen for these experiments because it is inhabited solely by Northern Pike, is relatively shallow (maximum depth < 6.1 m), and is easily accessible. Derk’s Lake stratifies seasonally, with a thermocline at approximately 4-m depth and anoxic conditions at greater depths (Table 1). Experimental design.—During June 2011, 111 Northern Pike were captured by angling in water less than 2 m deep at Alexander Slough (Susitna River drainage), located 24.1 km due west of Anchorage on the Kenai Peninsula. Fish were transported via floatplane to Derk’s Lake under permit from the Alaska Department of Fish and Game (ADFG) and with approval from the Animal Care and Use Committee at Montana State University. It was necessary to bring Northern Pike to the study site because the population in the lake had previously been suppressed by gillnetting. To minimize fish stress during transport, Northern Pike were maintained in two aerated, 1.2-m3 fish totes. Upon arrival at the TABLE 1. Ambient water conditions during randomized exposure treatments of Northern Pike from the Susitna River drainage, Alaska, to the 5,620.8-cm3 or 1,966.4-cm3 water gun on June 15, 2011 (SpC = specific conductance; DO = dissolved oxygen concentration).

Depth (m) 0 1 2 3 4 5 6

Temperature (◦ C)

SpC (S/m)

DO (mg/L)

pH

14.72 14.58 13.29 12.06 10.77 6.90 6.14

0.038 0.038 0.038 0.038 0.040 0.088 0.167

8.81 8.82 8.27 7.22 3.05 0.28 0.55

6.72 6.68 6.66 6.75 6.89 7.58 7.85

WATER GUN USE TO CONTROL NORTHERN PIKE

lake, the fish were transferred into 0.9- × 1.5- × 3.1-m holding pens constructed of 6.3-cm polyvinyl chloride (PVC) pipe with a 15.2-cm-mesh net and a 0.6- × 0.6-m flap at the top for access. Holes were drilled into the PVC frame to allow the holding pen to take on water and sink while being suspended below the lake surface by buoys. After placement into holding pens, fish were given a 24-h acclimation period prior to additional handling. All study fish were removed from the lake and placed into three aerated fish totes prior to anesthesia for evaluation of health and normal swimming behavior. Northern Pike were anesthetized with Tricaine S (Western Chemical, Ferndale, Washington) at a concentration of 100 mg/L of water; to prevent overcrowding, no more than 10 fish were placed in the anesthesia tank at one time. Fish remained in the anesthetizing tank until stage-4 anesthesia was reached (i.e., total loss of swimming motion, with weak opercular movement, as described by Yoshikawa et al. 1988). Individual fish were removed from the tank, measured (nearest 1 mm TL), and injected with a PIT tag (Biomark, Boise, Idaho) in the posterior region of the dorsal musculature. Subsequently, fish were placed in an aerated, 1.2-m3 recovery tank and were monitored for recovery. Signs of recovery included proper orientation, response to stimulus, and normal opercular movements. After a fish had completely recovered, it was placed into a holding pen (0.9 × 1.5 × 3.1 m) and allowed to acclimate in the lake for an additional 24 h prior to being used in experimentation. This acclimation period was provided to ensure that experimental mortality was not induced by stress from the tagging procedures. All 111 fish survived the tagging process.

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Water gun deployment.—Two water guns were used for this study: a Bolt Model 1900 prototype with a 5,620.8-cm3 (343-in3) chamber (Bolt Technology, Inc., Norwalk, Connecticut); and a Model S80 with a 1,966.4-cm3 (120-in3) chamber (U.S. Seismic Systems, Inc., Houston, Texas). During testing, a water gun was suspended from a buoy and jib crane atop a plastic pontoon float (2.4 m wide × 3.7 m long) in 3.7 m of water. A line was set in place by fastening a rope from parallel sides of the pontoon to each opposite shoreline so that the pontoon and gun maintained the same orientation and depth throughout the experiment (Figure 1). The water gun was positioned in a horizontal plane parallel with the lake surface, with the gun port facing away from the closest shoreline and the pontoon ensuring a constant linear depth profile of 3.7 m from surface to substrate. For each experiment, the water gun was lowered to a depth of 2.4 m. Distances of 3, 6, and 9 m from the port end of the water gun (the side where water is expelled), on the static line, were measured and marked using buoys to ensure consistent deployment of exposure cages and the high-pressure sensor during experimentation. Each water gun was operated manually by using a FastFiring Par Air Gun Firing Circuit (FC 100; Bolt Technology). Both of the water gun experiments were conducted at an operating pressure of 13.8 MPa (2,000 pounds per square inch [PSI]), and the pressure was maintained throughout the experiment by a 34.0 m3/h, 22-hp, 20.7-MPa (3,000-PSI)-rated electric compressor and a three-phase, 50-kW generator. Pulse pressure exposure.—Each water gun was tested separately. Northern Pike were individually exposed to the 5,620.8-cm3 water gun or the 1,966.4-cm3 water gun. To ensure

FIGURE 1. Photo of the experimental setup, including the pontoon barge with the water gun (in the water) suspended by the jib crane; also shown is an aluminum boat with a fish tote and the retrieval of an exposure cage used for Northern Pike. [Figure available online in color.]

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that the study fish did not receive any energy emitted from the water gun while they were submerged in the lake, Northern Pike were removed from the lake holding pens and were placed into four aerated, 1.2-m3 fish totes on land. Water temperature and water level in the totes were maintained with a sump pump that extracted lake water from 1–2-m depth, and all fish were maintained in the totes for an equal length of time during the course of the experiment. A single Northern Pike was then captured with a dip net, identified from its PIT tag, and placed into an exposure cage. A randomized design was used to assign exposure treatments (distance from the 5,620.8-cm3 water gun) of control, 3, 6, and 9 m to each individual fish (experimental unit). There were 15 replicates per treatment group (n = 15 replicates of one fish per exposure), for a total of 60 fish. To avoid potential confounding effects associated with fish size and response to sound pressure (Yelverton et al. 1975; Gaspin 1976; Lewis 1996), only individuals that fell within approximately 1 SD of the mean length (50.9 ± 12.6 cm) were used for the experiments (87 of the 111 available Northern Pike). To transport fish from shore to a predetermined distance marker, exposure cages containing the fish were placed into fish totes filled with lake water and were secured inside an aluminum boat. Water quality in the transport totes was actively monitored. Water in the totes was replaced after every 4–5 replicates since an individual fish would only reside in the transport tote for 1–2 min. Exposure cages consisted of a 0.9-m3 frame constructed of 5.1-cm PVC pipe and a 1.3-cm2-mesh net that was secured to the frame by plastic zip ties. Holes were drilled into the PVC to allow the cages to sink quickly and to allow trapped air to escape the pipe. Each of the exposure cages was removed from the tote and immediately lowered into the lake at the predetermined exposure distance, in line from the port end of the water gun, so that the center of the cage was set at a depth of 2.4 m from the surface. Due to the logistics of this study and the need to minimize the length of time for which fish were held in totes out of the lake, the fish were not acclimated in the exposure cages. In addition, fish buoyancy was not evaluated once the fish in the exposure cages were lowered into the lake. The cages were lowered to 2.4 m, which is only 1 m deeper than the pens in which fish were maintained for the duration of the study. Thus, the potential pressure change would be significantly less than an atmospheric pressure. A Tourmaline ICP underwater blast sensor (sensitivity = 0.73 mV/kPa; PCB Piezotronics, Inc., Depew, New York) was positioned to 2.4 m and suspended immediately in front of each exposure cage (between the water gun and the cage). After the fish exposure cages and blast sensors had been lowered into the water and after the boats had been positioned at a safe operating distance (>50 m), the water gun was fired twice at an operating pressure of 13.8 MPa (2,000 PSI). Control fish were treated in the same manner as treatment fish and were submerged to a depth of 2.4 m for less than 1 min at a point between 3 and 9 m from the water gun; however, the water gun was not discharged.

A feasibility experiment had previously established that a single exposure would not induce sufficient variability to establish a cumulative sound exposure level (SELcum ) or a continuum of responses or to establish a dose–response study with varying distances. After the two pulses were emitted, the exposure cages were immediately retrieved and placed in the fish tote on the boat. The fish were then ferried back to the shore and were returned to an aerated recovery tote for postexposure monitoring. The entire exposure procedure (i.e., netting of a single fish from a tote, scanning the PIT tag, placement in the exposure cage, transport to the predetermined distance and submersion, exposure to two pulses from the water gun, retrieval from the lake and transport back to shore, and placement in recovery totes) took less than 5 min. None of the fish was held out of water for more than 15 s during the entire process. At the completion of all exposures from the 5,620.8-cm3 water gun, Northern Pike were removed from the recovery totes and were placed in submerged holding pens within Derk’s Lake. Holding pens were suspended at the water’s surface by buoys and were moored to a floating dock in less than 3 m of lake water. After completion of the experiment, water temperature, conductivity, dissolved oxygen concentration, and pH were measured at 1-m intervals from the surface to 6-m depth (Table 1; these data were measured and analyzed by ADFG, Soldotna). The holding pens were monitored for mortalities every 24 h for a total postexposure period of 168 h (7 d). Fish were not fed during this holding period. Deceased fish were immediately removed and placed in a walk-in refrigerator within 30 min, and they were maintained at 6.7◦ C for 24 h prior to being necropsied (see below). No body decomposition was noticed in any of the fish that were removed from the pens at the scheduled 24-h monitoring periods; this was likely a function of the cold water temperature. At 168 h postexposure, all remaining fish were euthanized by prolonged exposure to Tricaine S and were stored in a refrigerator for 24 h prior to necropsy. The pulse pressure experiment was repeated with the 1,966.4-cm3 water gun, and fish were exposed to two pulse pressures at a distance of 3 or 6 m. Twenty-one Northern Pike (n = 7 fish/treatment, including the control) were selected from among the remaining 27 fish. Again, to avoid potential confounding, only fish within 1 SD of the mean length (49.6 ± 9.4 cm) were included in this experiment. After treatments were applied, all Northern Pike that were exposed to the 1,966.4-cm3 water gun were immediately euthanized by prolonged exposure to Tricaine S and were necropsied. There were not enough remaining days in the field to permit the 168-h monitoring of fish exposed to the 1,966.4-cm3 water gun; thus, statistical comparisons between the two experiments were not possible. The remaining Northern Pike that were not used in either experiment were also euthanized by prolonged exposure to Tricaine S. Necropsy and assessment.—After the Northern Pike had been refrigerated for 24 h, they were necropsied (Gaikowski 2003). A single incision was made on the ventral surface of each fish, starting at the vent, moving anteriorly, and ending at the

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TABLE 2. Number of Northern Pike in each category of the explosion damage criterion (EDC; a metric quantifying total internal organ damage) for experiments with the 5,620.8-cm3 and 1,966.4-cm3 water guns (EDC categories: 0 = intact fish, with no hematoma or rupture of any internal organs; 1 = hematoma in organs, but gas bladder is intact; 2 = ruptured organs and hematoma in organs, but gas bladder is intact; 3 = ruptured gas bladder; 4 = ruptured gas bladder and hematoma in other organs; 5 = ruptured gas bladder and other ruptured organs). The EDC categories of 0–2 indicate that the fish will survive, and EDC categories of 3–5 indicate fish mortality.

EDC category Distance treatment

0

1

2

3

4

5

Total

3m 6m 9m Control

0 0 1 11

0 0 0 4

5,620.8-cm3 water gun 0 0 1 0

0 0 1 0

1 3 5 0

14 12 7 0

15 15 15 15

0 0 0

1,966.4-cm3 water gun 0 0 0 1 0 0

3 2 0

3 2 0

7 7 7

3m 6m Control

1 2 7

pericardium. Great care was used to ensure that the internal organs of the peritoneum were not damaged by the blade. Gross external and internal damage was evaluated; however, in order to form a single metric that combined internal organ damage measures (Yelverton et al. 1975, Yelverton 1996; Lewis 1996), we only scored tissue damage to the gas bladder, kidney, and liver (i.e., damage present or absent) based on a modified explosion damage criterion (EDC) for tissue condition (i.e., intact, hematoma, or ruptured; Table 2). Feasibility studies conducted on Snake River Yellowstone Cutthroat Trout O. clarkii bouvieri and Northern Pike (J. A. Gross et al., unpublished data) suggested that injuries to the gas bladder, kidney, and liver (particularly gas bladder rupture) were good predictors of fish mortality; those observations were similar to previous findings of fish damage (associated with fish size) from explosives (Yelverton et al. 1975; Gaspin et al. 1976; Lewis 1996). A thorough necropsy can take 15–30 min per fish, so it was impracticable to evaluate other important tissues (e.g., brain trauma) in large numbers of fish. All necropsies were conducted blind to treatment, as fish identification was determined after necropsy was performed. Although multiple individuals participated in the fish necropsies, the lead scientist confirmed all tissue conditions. Acoustic data acquisition.—A blast pressure sensor measured sound pressure levels (SPLs) associated with each exposure. A 482C05 Signal Conditioner (PCB Piezotronics) was adjusted to 10 mA of current to enhance the signal strength because there was 180 m of cable from the high-pressure blast sensor. A National Instruments CompactDAQ Data Acquisition System (NI 9234 Accelerometer, with a signal input range of 5 to −5 V; National Instruments Corp., Austin, Texas) was attached to the data acquisition hardware. The digitized waveform produced by the water gun was recorded using LABVIEW Signal Express 2010 (National Instruments) on a Dell Latitude E6410 laptop computer.

The National Instruments CompactDAQ was calibrated prior to sound level determinations. A Protek 2-MHz Sweep/Function Generator (Model B8011; Protek Test and Measurement, Norwood, New Jersey) was connected to the CompactDAQ with a PCB Piezotronics Model 401B04 ICP Sensor Simulator. The sweep/function generator was used in place of the high-pressure sensor to calibrate the ICP sensor signal conditioner, conduct system verifications, and assess frequency responses. After all acceptance test procedures were finalized, the function generator and sensor simulator were removed, and the high-pressure blast sensor was reattached. Numerous test shots were taken prior to experimentation to ensure that the acquisition system, the water guns, and the ancillary equipment were operating properly. If work was temporarily halted, this verification process was repeated prior to additional experimentation. Software recorded the peak SPL (SPLpeak ) and peak-to-peak SPL (SPLp-p ) in an amplitude spectrum and frequency. The SPLpeak (V) was the highest decibel level experienced by a fish, and the SPLp-p (V) was the voltage range between the highest and lowest pressures. The amplitude spectrum (dB) was recorded on a 0–25-kHz scale. For each fish exposure, the data acquisition system was set to record for 5 s at a rate of 51.2 kHz, with 256,000 samples recorded (51,000 measurements/s). The 5-s recording interval ensured that background levels prior to and after the signal were recorded. The actual source signal measured from the water gun was less than 100 ms of the 5 s that were recorded for each water gun pulse. The SELcum was derived from the sum of the two pulses recorded by the high-pressure sensor near the exposure cage by integrating the square of each SPLpeak after the signal breached 10 mV for 70 ms (Stadler and Woodbury 2009). The actual SELcum was assessed from 3,584 sample points (mV) recorded for each individual pulse, or 7,168 measurements recorded for the sum of the two pulses for each exposed fish.

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Data analysis.—Logistic regression was used to evaluate whether the probability of mortality at 24, 72, and 168 h postexposure (cumulative mortality) was related to the received SPL. Logistic regression relates the probability of mortality (π) to sound pressure variables through the logit link as follows: logit(π/[1−π]) = β0 + β1 (sound pressure) + β2 (fish length). In logistic regression, the estimated effect of a one-unit increase in sound pressure is a (exp[β1 ]) multiplicative change on the odds of mortality after accounting for fish length (R Development Core Team 2011). Tissue damage (as evaluated by the EDC) was analyzed with cumulative logistic regression (Christensen 2012) because the response variable (EDC) is an ordered multinomial variable. In cumulative logistic regression, the logit link function connects the cumulative probabilities, specifically the probability of being in EDC category j or lower, to sound pressure meaP(Yi ≤ j) ] (Agresti 2010). surements: logit[P(Yi ≤ j)] = loge [ 1−P(Y i ≤ j) The estimated effect of sound pressure is the same for each cumulative probability; therefore, the multiplicative effect on the odds of moving into a higher EDC category is the same for a one-unit change in sound pressure. Three pressure measurements (SPLpeak , SPLp-p , and SELcum ) were determined each time the water gun was discharged, and the mean of measurements associated with the two discharges was estimated for each fish. For the 5,620.8-cm3 water gun, all three measures were highly correlated (r > 0.80). However, for the 1,966.8-cm3 water gun, only SPLpeak and SPLp-p were highly correlated (r = 0.991); therefore, results for SPLpeak and SELcum are reported for both water guns. To avoid the problem of multicollinearity (poor estimation of model coefficients and associated SEs), each pressure measurement was included individually in the statistical models. The experimental methods were designed to minimize the possible confounding of fish length in the evaluation of mortality as a response to sound pressure. However, to be conservative, we included fish length as a covariate in the statistical models, although it is unlikely to be statistically important. Therefore, the effect of pressure was modeled after accounting for any minor differences in fish length. In the 5,620.8-cm3 water gun experiment, the pulse pressure data for the first 16 fish (of the total n = 60 fish) were either missing or substantially skewed because of technical issues with the high-pressure blast sensor. During exposure trials with the 1,966.4-cm3 water gun, software errors prevented the recording of the second pulse signal. As a result, some of the pulse pressure data were missing for the second experiment (7 of the 21 fish) as well. Discarding all cases with missing data would have resulted in a tremendous loss of information and wasted effort and expense; therefore, a multiple imputation procedure was used to statistically “fill in” the missing values. Missing data in this study could be managed because treatments (3, 6, and 9 m and the control) were randomly assigned to experimental units (i.e., missing completely at random). Furthermore, because the or-

der of treatments was randomized, sensor malfunction occurred irrespective of treatment group. Multiple imputation was used to recover data on pressure and time. The SPLpeak , SPLp-p , and SELcum exhibited essentially normal distributions within each distance class (3, 6, and 9 m and the control). Thus, multivariate normal values were imputed by using the mean and SD of the observed distribution within each group. Missing values for elapsed time between pulses were imputed using the mean of the observed distribution as the rate parameter of a Poisson distribution—the traditional method of modeling the period between the two pulses (Casella and Berger 2002). The imputed values were consistent with the observed values in terms of variation and mean value at each distance for each of the water guns (Tables 3, 4). Imputation procedures created multiple values for a single missing value, resulting in 10 data sets for SPLpeak , SPLp-p , and SELcum . Models were fit to each imputed data set, and the 10 parameter estimates and SEs were averaged (see Rubin 1987 for theoretical basis). For example, a regression coefficient for X 1 (for 10 imputations, denoted by j) would be estimated as 1 !10 ˆ βˆ 1 = 10 j=1 β1 j . Hypothesis tests were based on a normal approximation. RESULTS Received Pressures versus Distance from a Water Gun For both water guns, there was a clear separation of received SPLpeak between distance classes based on completely observed data; although there was some overlap between the 6- and 9-m treatment groups, differences in the received mean SPLpeak were statistically significant (ANOVA: F 3, 43 = 271.6, P < 0.0001 for observed data from the 5,620.8-cm3 water gun; F 2, 18 = 113.4, P < 0.0001 for observed data from the 1,966.4-cm3 water gun). Differences in average SELcum among distance classes were also statistically significant based on observed data (Kruskal– Wallis rank-sum test: χ2 = 54.6, df = 3, P < 0.0001 for the 5,620.8-cm3 water gun, Table 3; χ2 = 17.9, df = 2, P ≤ 0.0001 for the 1,966.4-cm3 water gun, Table 4). The clear distinction between mean pressure values for the different distance classes further supported the use of multiple imputation for the unobserved pressure values based on distance class. Each fish received two pulses from a water gun, with a median recovery interval between pulses of 23 s (range = 17–37 s) from the 5,620.8-cm3 water gun and 25 s (range = 19–121 s) from the 1,966.4-cm3 water gun. 24-h Cumulative Mortality from the 5,620.8-cm3 Water Gun Increasing SPLpeak increased 24-h cumulative mortality of Northern Pike (z = 2.077, P = 0.0378) after accounting for fish length and elapsed time between pulses from the 5,620.8cm3 water gun. However, there was no evidence that elapsed

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TABLE 3. Comparison of observed and imputed distributions of variables for the 5,620.8-cm3 water gun (SPLpeak = peak sound pressure level; SPLp-p = peak-to-peak sound pressure level; SELcum = cumulative sound exposure level). Estimates are the mean ± SE, with observed (OBS) or imputed (multiple imputation [MI]) sample sizes in parentheses. Imputed sample sizes are multiples of 10, since 10 random normal values (or Poisson values for time) were generated for each missing value.

Distance class Pressure variable SPLpeak (kPa), OBS SPLpeak (kPa), MI SPLp-p (kPa), OBS SPLp-p (kPa), MI SELcum (dB), OBS SELcum (dB), MI Elapsed time (s), OBS Elapsed time (s), MI

3m 1,132.2 1,175.6 1,329.7 1,321.3 204.8 204.7 22.0 21.8

± ± ± ± ± ± ± ±

6m

192.2 (18) 162.8 (12 × 10) 109.8 (17) 136.1 (13 × 10) 1.1 (10) 1.1 (5 × 10) 5.0 (8) 4.9 (7 × 10)

618.2 628 780.7 777.3 201.5 201.5 25.4 26.1

time between pulses was associated with 24-h cumulative mortality after accounting for fish length and SPLpeak (z = 1.546, P = 0.1222). There was also no evidence of an association between 24-h cumulative mortality and SELcum (z = 1.141, P = 0.2538) after accounting for fish length. Nine Northern Pike were removed as mortalities from the holding pens at 24 h postexposure. 72-h Cumulative Mortality from the 5,620.8-cm3 Water Gun There was an increase in 72-h mortality of Northern Pike with an increase in received SPLpeak after controlling for fish length (z = 3.109, P = 0.0019; Figure 2a). The estimated probability of mortality by 72 h for a fish receiving 1,379.0 kPa (200 PSI) of SPLpeak was 7.17 times greater than the probability of mortality for a fish receiving 689.5 kPa (100 PSI; 95% confidence interval = 2.07–24.89 times greater). The estimated probability of mortality was 0.282 for a 51.5-cm fish (mean length) receiving 689.5 kPa (100 PSI) of SPLpeak and was 0.738 for a fish

± ± ± ± ± ± ± ±

84.2 (20) 86.9 (10 × 10) 102.7 (19) 131.9 (11 × 10) 1.1 (10) 1.1 (5 × 10) 6.2 (9) 5.3 (6 × 10)

9m 432.9 436.9 596 608.5 199.5 199.6 25.4 24.8

± ± ± ± ± ± ± ±

44.8 (24) 46.3 (6 × 10) 46.3 (24) 49.3 (6 × 10) 0.4 (12) 0.3 (3 × 10) 6.2 (12) 5.3 (3 × 10)

receiving 1,379.0 kPa (200 PSI) of mean SPLpeak (Figure 2a). Similar to 24-h cumulative mortality, there was no evidence of an effect of elapsed time between pulses on 72-h mortality after accounting for fish length and the received SPLpeak (z = 0.9587, P = 0.3379). By 72 h postexposure, 16 fish (cumulative total) had been found dead and removed from the holding pens, with six of the seven new mortalities removed at 48 h postexposure. Unlike 24-h mortality, there was evidence that 72-h cumulative mortality had a positive association with SELcum (z = 2.223, P = 0.0263; Figure 2b) after accounting for fish length. The estimated probability of mortality was 0.202 for a 51.5-cm fish (mean length) receiving 200 dB of SELcum and was 0.678 for a fish receiving 206 dB of SELcum . 168-h Cumulative Mortality from the 5,620.8-cm3 Water Gun The mortality data for 168 h postexposure were identical to the results for 72 h postexposure. Mortality at 72 h and at

TABLE 4. Comparison of observed and imputed distributions of variables for the 1,966.4-cm3 water gun (SPLpeak = peak sound pressure level; SPLp-p = peak-to-peak sound pressure level; SELcum = cumulative sound exposure level). Estimates are the mean ± SE, with observed (OBS) or imputed (multiple imputation [MI]) sample sizes in parentheses. Imputed sample sizes are multiples of 10, since 10 random normal values (or Poisson values for time) were generated for each missing value.

Distance class Pressure variable SPLpeak (kPa), OBS SPLpeak (kPa), MI SPLp-p (kPa), OBS SPLp-p (kPa), MI SELcum (dB), OBS SELcum (dB), MI Elapsed time (s), OBS Elapsed time (s), MI

3m 592.2 584.4 725.8 731.1 198.7

101.6 (12) 88.8 (2 × 10) 102.6 (12) 105.6 (2 × 10) 1.5 (7) NA (0) 59.6 ± 52.5 (5) 60.6 ± 9.2 (2 × 10) ± ± ± ± ±

6m 367.6 351.9 506.7 493.8 194.6 195

± 78.9 (11) ± 79.2 (3 × 10) ± 56.9 (7) ± 65.4 (7 × 10) ± 1.7 (4) ± 1.7 (3 × 10) 21.0 (1) 21.4 ± 4.3 (6 × 10)

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FIGURE 2. Fitted probabilities from logistic regression models to multiply imputed data sets for 72-h mortality for a Northern Pike of mean length (51.5 cm) across (a) the observed range of peak sound pressure level (SPLpeak ) values (kPa) and (b) the experimental range of observed cumulative sound exposure level (SELcum ) values (dB) for the 1,966.4-cm3 water gun. Confidence band for each regression line is a 95% Workman–Hotelling band for the correct mean response within the entire range of pressure values. Proportions of observed mortality (P[mortality]) are plotted with the fitted curve. Each point represents a distance class (control or 3, 6, or 9 m), the proportion of mortality in that group, and the mean SPLpeak or mean SELcum for that group.

168 h occurred when SELcum increased above 195 dB. The minimum SELcum calculated for gas bladder rupture resulting from exposure to the 5,620.8-cm3 water gun at a distance of 9 m was 199 dB referenced to 1 µPa2/s.

Explosion Damage Criterion and Internal Organ Damage The majority of fish that were exposed to sound energy from the 5,620.8-cm3 water gun were classified as having ruptured gas bladders and other hemorrhaging organs (EDC category 4) or as having ruptured gas bladders and other ruptured organs (EDC category 5; Table 2). For the 5,620.8-cm3 water gun, mortality by 168 h postexposure was observed only in fish with an EDC score of 5. There was consistently more tissue damage in fish receiving pulse pressure than in control fish. For example, fish in the control group for the 1,966.4-cm3 water gun experiment suffered no damage (Table 2). Similarly, for the experiment with the 5,620.8-cm3 gun, minor damage was documented in fish that were randomly assigned to the control group (Table 2). Fish that were located 3 m from a water gun suffered more tissue damage than fish that were situated further from the water gun (Table 2). In general, greater internal damage (higher EDC category) was associated with a greater SPLpeak from both the 5,620.8cm3 and 1,966.4-cm3 water guns after accounting for fish length (z = 4.833, P ≤ 0.0001 for the 5,620.8-cm3 gun; z = 2.772, P = 0.0056 for the 1,966.4-cm3 gun). The probability of internal damage consisting of a ruptured gas bladder and other organ damage (EDC category 3 or higher) increased with an increase in received SPLpeak (Figure 3a, b, line labeled with EDC ≥ 3). Correspondingly, the probability of having all organs intact was highest for fish with low levels of received SPLpeak (Figure 3a, b, line labeled with EDC = 0). The minimum calculated SELcum for gas bladder rupture after exposure to pulses from the 1,966.4-cm3 water gun at a distance of 6 m was 194 dB referenced to 1 µPa2/s. The SELcum was not associated with the probability of a higher EDC category for fish exposed to the 1,966.4-cm3 water gun (z = 0.511, P = 0.6097), but it was positively associated with an increase in EDC category for fish exposed to the 5,620.8-cm3 water gun (z = 4.506, P ≤ 0.0001). For the 5,620.8-cm3 water gun exposures, there was evidence that the probability of a higher EDC category increased with an increase in elapsed time between pulses (z = 2.177, P = 0.0298). With the 1,966.4-cm3 water gun, however, the relationship between EDC category and elapsed time between pulses was not statistically significant (z = –0.654, P = 0.5131). DISCUSSION This study provides what we believe to be the first comprehensive investigation into the influence of pulse pressure from water guns on the mortality of aquatic organisms. Water guns had been previously tested to evaluate hearing loss in marine mammals (Finneran 2002), to alter eel passage in hydropower facilities (P. Chelminski, Bolt Technology, unpublished data), and to replicate a gaping response in oysters after pulse pressure exposure from a sparker and an air gun (Paparella and Allen 1970; P. Chelminski, Bolt Technology, unpublished data). We evaluated the potential of pulse pressure emitted from two types


FIGURE 3. Fitted probabilities from cumulative logistic regression models to multiply imputed data sets for explosion damage criterion (EDC) data for a Northern Pike of mean length (51.5 cm) across the observed range of peak sound pressure level (SPLpeak ) values (kPa) for (a) the 5,620.8-cm3 water gun and (b) the 1,966.4-cm3 water gun. The P(EDC) is the probability of a fish being assigned to EDC categories of 0, 1–2, or ≥3, based on the degree of gross internal organ damage (see Table 2 for definition of EDC categories).

of water guns to suppress and deter the expansion of invasive Northern Pike, and we determined the SELcum required to induce mortality associated with soft tissue injury. The 1,966.4-cm3 water gun caused a similar number of gas bladder ruptures as the 5,620.8-cm3 water gun, but it produced approximately half the mean SPLpeak at each distance. With the 1,966.4-cm3 water gun, 100% of fish in the 3- and 6-m distance treatments exhibited rupture of the gas bladder. The mean SPLpeak was 367.6 kPa (53.3 PSI) at 6 m from the 1,966.4-cm3 water gun. The mean

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SPLpeak at 9 m from the larger water gun was 449.4 kPa, as gas bladders were ruptured in 13 of the 15 Northern Pike in that treatment. This represents the first experiment in which fish have been exposed to energy from a 1,966.4-cm3 water gun. In 2010, we conducted a feasibility study at the exact same lake and location to evaluate the potential effects of water guns on Northern Pike, and we utilized a 5,620.8-cm3 water gun for that study (J. A. Gross, unpublished data). Although the 1,966.4-cm3 water gun is much smaller, it induced a similar degree of tissue insult as recorded from the 5,620.8-cm3 water gun. The position of a fish in relation to the source (e.g., closest to or farthest away from the source) may affect the outcome on other fish because an individual that is nearest to the source may provide a shield for fish that are farther away. In the 2010 feasibility study, three fish (TL range = 34.7–60.0 cm) were placed in each exposure cage (n = 5 cages, with 3 fish/cage). Four (26.6%) of the 15 fish had ruptured gas bladders. This finding would be important to consider if pulse pressure is to be used as a barrier to fish movement because the fish nearest to the source would receive the most damaging energy. A fish that is farthest from the source could conceivably swim past a barrier—or if stunned, be pushed through the barrier with water current—between pulses. This type of barrier would be most effective for low-flow situations in which (1) firing intervals from a single energy source could be significantly increased or (2) multiple water guns are fired quickly while staggering the water gun configuration. To establish a no-passage or lethal-pulse pressure barrier, the firing interval would have to be shortened between pulses but not at the cost of decreasing the maximum output pressure. In this study, increasing the number of pulse pressures and SELcum increased the likelihood of soft tissue damage. Although this may seem intuitive, the equal-energy hypothesis suggests that the effects of a large single pulse of energy are equivalent to the effects of energy received from many smaller pulses (Smith and Gilley 2008). Although our study was not designed to evaluate that hypothesis, previous studies on the effects of pulse pressure associated with pile driving do not support the hypothesis for tissue damage (i.e., fewer louder strikes were more effective than many quieter strikes; Halvorsen et al. 2012). It is currently hypothesized that the exposure window for fish to suffer no effect from previously received energy is 12 h (Stadler and Woodbury 2009). In the present study, Northern Pike were exposed to two pulses from a water gun. Although statistically we saw an effect of time (i.e., an increased probability of a higher EDC category with an increase in elapsed time between pulses) for the larger water gun, the difference in median recovery interval between pulses for the larger water gun was only 2 s. Anecdotally, the first pulse tended to stun or disorient the Northern Pike, as fish would sink or list to one side, and the second pulse pressure caused fish stunning and subsequent settling at the bottom of the exposure cage. Once the fish were removed from the exposure cage and placed in a recovery tank, they took up to 1 h to gain

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equilibrium. This is most likely a function of SELcum , as the degree of stunning decreased with distance from the source. The present study was focused on determining lethal thresholds to suppress invasive Northern Pike. We assume that the probability of mortality would have increased after 14–21 d, as only 31% of the fish were dead by 168 h (7 d) postexposure despite the fact that almost 96% of all fish had severely damaged gas bladders and significant soft tissue damage, especially in the kidney. Fish have been documented to recover from gas bladder injury. For example, Bellgraph et al. (2008) reported that juvenile Rainbow Trout O. mykiss could survive a surgical rupturing of the gas bladder and that the injury was completely repaired after 14 d. Those authors had hypothesized that gas bladder rupture would result in direct mortality, but the study results refuted this hypothesis, showing minimal mortality in fish experiencing barotrauma from surgical ruptures. The important differences

between the Bellgraph et al. (2008) study and our study were the size of the fish, the thickness of the gas bladder, and the relative uniformity and small size of surgically created ruptures. Gas bladder rupture caused by pulse pressure exposure from the larger water gun was characterized by large, non-uniform cavities in the anterior and posterior portions of the gas bladder. Ruptures associated with the 1,966.4-cm3 water gun were smaller, but we did not quantify the magnitude of the rupture and we did not take photos of every fish. It would be difficult to imagine that Northern Pike could heal from the injuries photographed in fish that survived through day 7 (Figure 4). Bellgraph et al. (2008) suggested that the probability of additional mortality increases with additional injuries from barotrauma. Gas bladder rupture in the present study was associated with tissue damage in other organs near the rupture, and such injuries would subsequently lead to additional mortality (Gaspin 1976). Other factors, such as gas bladder volume, can directly alter the risk of barotrauma.

FIGURE 4. Photos of common injuries in Northern Pike that were exposed to pulse pressure from the 5,620.8-cm3 water gun. The leftmost image is of a control fish (L = liver; GB = gas bladder; S = stomach; RGB = ruptured gas bladder; G = gonad; RK = ruptured kidney). [Figure available online in color.]


When exposed to pulse pressure, a negatively buoyant fish (i.e., a fish with a deflated gas bladder) could experience less trauma (Stephenson et al. 2010). In the present study, reported values would be conservative, underestimating a response if fish were negatively buoyant. Tissue damage, specifically hematomas in the liver and kidney, was recorded for a small number of fish in the control group (Table 2). Because these fish were not exposed to energy from the water gun, damage was likely an artifact of fish handling and transport. All dead fish were placed in large trash bags and transported by truck to a refrigerator for necropsy 24 h later. Because the numbers of euthanized fish were greater at the completion of the study (∼70% survival by 168 h), some of the hematomas in soft tissue could have been related to the weight of fish stored together in transport or to leakage of blood vessels during refrigeration. It should be noted that no control fish presented a ruptured organ or ruptured gas bladder. Water guns may be an effective tool for suppressing invasive Northern Pike along with other nuisance fishes, such as Northern Snakeheads Channa argus, that occupy similar niches and habitat types. Additional research is needed to evaluate the potential for deployment of water guns as a static permanent or temporary barrier, either stationary or mobile, and for operating water guns at lethal or nonlethal levels. Northern Pike may be especially susceptible to water guns, as they are territorial and exist in particularly defined water depths and habitats (Inskip 1982; Haught and von Hippel 2011). An integrated suppression approach combining a variety of modalities, such as water guns, gillnetting, electricity, and even limited chemical application in shallow waters, may lead to effective Northern Pike suppression. These water guns do have limitations, such as initial costs, size, shallow submeter water depth operation, and possible human health hazards associated with high-pressure air requirements. Additionally, when present, nontarget species could also be impacted behaviorally and physiologically. Sound energy does offer significant advantages, however, because once discharged, the energy and sound are quickly attenuated, and pulse pressure from a single water gun can be discharged every couple of seconds. The use of sound energy to suppress invasive fishes or to direct fish movement has been reviewed (Popper et al. 2005; Noatch and Suski 2012). Although the requirements for methods to eradicate fish and to prevent or direct fish passage are varied (e.g., hydropower exclusion of salmonids from water intakes; prevention of Asian carp passage from the Mississippi River to the Great Lakes through the Chicago Shipping and Sanitary Canal), water guns may be effective as a tool for fisheries management. ACKNOWLEDGMENTS This work was funded by the Alaska Sustainable Salmon Fund (Project Number 44613). We thank the ADFG Division of Commercial Fisheries, Soldotna, and the ADFG Division of Sport Fisheries, Palmer, for providing logistical and technical

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support in the field. We thank Nicholas Thompson for statistical support. We also thank Ken deFriesse (Bolt Technology, Inc.) for field and logistics support. Any use of trade, product, or firm names is for descriptive purposes only and does not imply endorsement by the U.S. Government. REFERENCES ADFG (Alaska Department of Fish and Game). 2002. Alaska aquatic nuisance species management plan. ADFG, Juneau. Agresti, A. 2010. Analysis of ordinal categorical data, 2nd edition. Wiley, New York. Baxter, C. V., K. D. Fausch, M. Murakami, and P. L. Chapman. 2004. Fish invasion restructures stream and forest food webs by interrupting reciprocal prey subsidies. Ecology 85:2656–2663. Bellgraph, B. J., R. S. Brown, J. R. Stephenson, A. E. Welch, K. A. Deters, and T. J. Carlson. 2008. Healing rate of swim bladders in Rainbow Trout. Transactions of the American Fisheries Society 137:1791–1794. Boucher, D. 2003. Illegal fish stockings threaten Maine lakes and rivers. Maine Department of Inland Fisheries and Wildlife, Rangeley Lakes Regional Headquarters, Strong. Available: www.maine.gov/ifw/fishing/ illegal stocking.htm. (August 2012). Byström, P., J. Karlsson, P. Nilsson, T. Van Kooten, J. Ask, and F. Olofsson. 2007. Substitution of top predators: effects of pike invasion in a subarctic lake. Freshwater Biology 52:1271–1280. Casella, G., and R. L. Berger. 2002. Statistical inference, 2nd edition. Duxbury, Pacific Grove, California. Christensen, R. H. B. 2012. Ordinal: regression models for ordinal data, R package version 2012.01-19. R Foundation for Statistical Computing, Vienna. Available: www.cran.r-project.org/package=ordinal./ (August 2012). Finneran, J. J., C. E. Schlundt, R. Dear, D. A. Carder, and S. H. Ridgway. 2002. Temporary shift in masked hearing thresholds in odontocetes after exposure to single underwater impulses from a seismic watergun. Journal of the Acoustical Society of America 111:2929–2940. Gaikowski, M. P., J. C. Wolf, S. M. Schleis, and W. H. Gingerich. 2003. Safety of oxytetracycline (Terramycin TM-100F) administered in feed to hybrid Striped Bass, Walleyes, and Yellow Perch. Journal of Aquatic Animal Health 15:274–286. Gaspin, J. B., M. L. Wiley, and G. B. Peters. 1976. Experimental investigations of the effects of underwater explosions on swimbladder fish, II. 1975 Chesapeake Bay tests. Naval Service Weapons Center, Technical Report NSWC-IH, Silver Spring, Maryland. Halvorsen, M. B., B. M. Casper, C. M. Woodley, T. J. Carlson, and A. N. Popper. 2012. Threshold for onset of injury in Chinook Salmon from exposure to impulsive pile driving sounds. PLoS (Public Library of Science) ONE [online serial] 7(6):e38968. DOI: 10.1371/journal.pone.0038968. Haught, S., and F. A. von Hippel. 2011. Invasive pike establishment in Cook Inlet basin lakes, Alaska: diet, native fish abundance and lake environment. Biological Invasions 13:2103–2114. Hutchinson, D. R., and R. S. Detrick. 1984. Water gun vs air gun: a comparison. Marine Geophysical Researches 6:295–310. Inskip, P. D. 1982. Habitat suitability index models: Northern Pike. U.S. Fish and Wildlife Service Biological Services Program FWS/OBS-82/10.17. Johnson, B. M., P. J. Martinez, J. A. Hawkins, and K. R. Bestgen. 2008. Ranking predatory threats by nonnative fishes in the Yampa River, Colorado, via bioenergetics modeling. North American Journal of Fisheries Management 28:1941–1953. Lewis, J. A. 1996. Effects of underwater explosions on life in the sea. Australian Department of Defence, DSTO (Defence Science and Technology Organisation) Aeronautical and Maritime Research Laboratory, Melbourne. McMahon, T. E., and D. H. Bennett. 1996. Walleye and Northern Pike: boost or bane to Northwest fisheries? Fisheries 21(8):6–13. Muhlfeld, C. C., D. H. Bennett, R. K. Steinhorst, B. Marotz, and M. Boyer. 2008. Using bioenergetics modeling to estimate consumption of native

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