THESIS PROPOSAL
Field Trials for the Use of Sea Lamprey Migratory Pheromone as an Attractant within Tributary Systems
Wayne R. Bouffard
RubensteinSchool of the Environment and Natural Resources
Aquatic Ecosystems and Watershed Sciences
April 2008
Literature Review
Taxonomy
Biology of the sea lamprey
Sea lamprey control
Integrated Pest Management
Pheromones
Chemoreception and fish
Pheromones and sea lamprey
Thesis Proposal
Statement of Problem
Objectives
Research Questions
Hypotheses
Methods
Study site
Pheromone collection
Passive Integrated Transponder (PIT Tags)
Field trials
Data analysis
Management Implications
Literature Cited
Thesis Proposal
Statement of Problem
The importance of a migratory pheromone has been well documented in the laboratory (Li et al. 1995, Bjerselius et al. 2000, Vrieze and Sorensen 2001) and results of limited field experiments are promising (Bjerselius et al 2000, Wagner et al. 2006)In order for migratory pheromone to be used for population control, its effectiveness under natural field conditions needs to be documented and quantified. The role of a migratory pheromone in guiding sea lamprey to specific areas within a tributary system has yet to be evaluated. This function would seem plausible, since many of the tributary systems used by sea lamprey are large and include smaller tributaries. Pheromones could also be effective at enhancing trap and removal operations intended for population control. The goal of this study is to evaluate the efficacy of migratory pheromone applications to redistribute migratory sea lamprey within a tributary system and enhance trapping effectiveness in the presence of background pheromone.
Objectives
- Determine whether migratory sea lamprey will move toward an artificially-increased larval pheromone concentration within a stream system in the presence of background levels of pheromone from stream-resident larvae.
- Determine whether larval sea lamprey pheromones can be used to effectivelyredistribute adult sea lampreyduring their migration.
Research Questions
1)Is there a change in proportion of tagged lamprey entering a tributary stream (Pond Brook) of Malletts Creekduring pheromone application (ON) vs no pheromone application (OFF)?
Response variables:
- Change in # tagged lamprey captured in Pond Brook
- Change in # wild lamprey captured in Pond Brook
2)Is there a change in the rate of movement of tagged lamprey during ON vs OFF?
3)Is the difference in # lamprey entering Pond Brook vs. Malletts Creek during pheromone application of management significance?
4)Is the total number of untagged lamprey caught in both streamsduring the season in which pheromones are applied higher than historic levels?
Hypotheses
1)πc= proportion of lamprey captured in Pond Brook without pheromone
πp= proportion of lamprey captured in Pond Brook with pheromone
H0: πc = πp
Ha: πp > πc
α = 0.05
2)τc= time to pass upstream antenna in Malletts without pheromone
τp=time to pass upstream antenna in Malletts with pheromone
H0: τc = τp
Ha: τp > τc
3)H0: πp < .70
Ha: πp ≥ .70
4)NT = Total number of lamprey captured
NH = Historical average of lamprey captured
H0: NT = NH
Ha: NT > NH
Methods
Study site
For a study site we will use Malletts Creek and its tributary Pond Brook in Colchester, VT. This is an ideal field site for a number of reasons. There have been ongoing adult sea lamprey trapping operations in both streams since 2000, conducted by the U.S. Fish and Wildlife Service. Both streams are small,enabling effective trapping of migrating sea lamprey using a temporary weir and portable assessment traps. Both streams also fall within the size limitations for effective use of Passive Integrated Tag (PIT) receivers to track the movements of tagged fish. Neither stream has been treated with lampricides as part of the sea lamprey control program. There is a population of sea lamprey larvae in Malletts Creek which will produce the background pheromone. Based on historical trapping data a very small percentage of sea lamprey choose to migrate up Pond Brook. The most recent survey of Pond Brook (2001) estimated the larval population to be 1,113; in Malletts Creek a 2005 survey estimated 2,442 larvae (USFWS unpublished data). The discharge of Pond Brook was calculated during November of 2006 to be approximately 12 cfs. This flow is most likely representative of workable flows during the spring spawning migration period. The discharge of Malletts creek was determined to be approximately 49 cfs at bankfull conditions which are representative of workable spring flows.
Pheromone collection
During the summer of 2006 approximately 2,500 sea lamprey larvae were collected using backpack electrofishing units from Putnam Creek in Crown Point, NY. They were transported to an 800L holding tank with approximately 8 cm depth of sand substrate at the University of Vermont’s Rubenstein Ecosystem Science Laboratory in Burlington, VT. The tank was supplied with a constant flow of dechlorinated water (at a rate of approximately 0.6 L/min) and fed weekly with a slurry of bakers yeast to achieve a concentration of 300 mg/L (Mallatt 1983, Rodríguez-Muñoz 2003). The overflow of the tank was diverted first through a 75 micron inline filter, then through a class column packed with XAD-7 high performance resin. Once a week, concurrent with feeding, flow through the column was stopped and the column was rinsed with 2.5L of methanol to release any organic molecules that had been adsorbed to the resin. This extract was stored at -20º C. This extract was then condensed using a rotary evaporator to reduce approximately 10 fold (1000ml extract to 100ml concentrated whole larval washings). These procedures were developed by the GLFC (unpublished protocol). Currently there is no accurate analysis developed for determining the exact concentration of the pheromone components, as such we will use larval equivalents for reporting results.
Passive Integrated Transponder (PIT Tags)
Passive Integrated Transponder(PIT tags) are small (23mm x 1mm) glass cylinders which are pre-programmed with a unique numeric code. These tags are passive in that there is no battery in the tag; they are activated when they are passed through or in close proximity to an antenna of the correct inductance and capacitance. Placement, survival, and retention of PIT tags in sea lamprey were evaluated during the winter of 2006-7. Twenty four parasitic phase lamprey, collected by local fisherman, were implanted with tags during the period 22 November 2006 - 11 January 2007. Tags were implanted by making a small incision in the skin using a sterile scalpel and inserting the tag. The initial 5 tags were placed to evaluate tagging location. Three were placed dorsally subcutaneously and were shed within 8 days, two were placed intraperitoneally and remained in the lamprey six and seven weeks until death of the animal. All of the remaining 19 tags were implanted intraperitoneally; only one surgery resulted in infection and subsequent tag loss. All other tags were retained throughout the period of observation (through February 2007) or until death of the animal, regardless of whether sutures were used to close the incision following implantation. There were a total of 4 mortalities during the observation period; however these are most likely attributable to starvation. The lamprey were collected as actively feeding parasites and held in laboratory conditions without hosts.
Field trials
Response of migratory lamprey will be evaluated in two ways: (1) PITtagged migratory phase sea lamprey will be released downstream from the confluence of Malletts Creek and Pond Brook to track their movements, decision-making behavior, and ultimate decision as to which stream to enter. (2)The number of migratory sea lamprey captured in each stream during the “on” and “off” phases will be compared to determine whether more are trapped in Pond Brook when the larval odor is being pumped.
Pre-spawning migratory sea lamprey will be collected by the US Fish and Wildlife Service during their annual trapping operations,implanted intraperitoneally with a PIT tag, transported to a holding cage in Malletts Creek,where they will be held for a minimum of 24 hours to fully recover from the stress of handling and surgery prior to being released in Malletts Creek during a trial. Continuously monitoring stationary PIT tag receivers will be set up at three sites: one in Malletts Creek above the confluence, one in Pond Brook, and one approximately 110m downstream of the confluence of the two streams to monitor their movement up Malletts Creek, into Pond Brook, or out of the study area,respectively (Fig 1.).
In streams less than 15 m wide and 1 m deep, a single antenna can detect lamprey passing the receiver at any point on a transect perpendicular to the flow. Two receivers may be used to determine direction of movement. The information coded in the tag permits recording of the movements of individually-identifiable lamprey and the time when it passed the antenna. Information from the receivers will be recorded continuously using a Destron FTS 2001 receiver (Oregon RFID) housed in a weather-proof box; data will be downloaded every 24 - 48 h. The functioning of the antennas will be checked every day by towing a PIT tag in the stream past each antennato ensure that the detectors are functioning properly.
Twenty lamprey will be tagged and released for each trial and traps will be checked daily. Our goal is to release at least 100 tagged lamprey below the confluence of the two streams. Adult capture data from previous years suggests that this is a realistic number. Because we wish to avoid holding lamprey prior to tagging and release, the number of animals released each day will vary; on days when fewer than 15 lamprey are available for tagging and release, none will be tagged. Trials will start after sunset. Concentrated whole larval washings will be metered into Pond Brook upstream of the trap using a peristaltic pump to achieve concentrations above those in Malletts Creek. Pheromone will be applied on alternating nights for treatment and control trials. Each trial will run for approximately 5 hours, beginning at or near 2200 hours and continuing until 0300 hours. This is the period of the evening when the majority of migratory movements occur (Kelso and Gardner 2000). Traps will be checked immediately prior to and following pheromone application. Tagged lamprey will be held in a release cage for one half hour while the pheromone is applied prior to release.
Portable assessment traps (PATs; 0.6m wide x 0.6m high x 1m long), similar to those described by Schuldt and Heinrich (1982) and Hunn and Youngs (1980) will be set up in
Malletts Creek and Pond Brook above the confluence of the two streams, using wings constructed of 13mm wire mesh to block lamprey from passing the traps. The use of these traps will serve as a backup mechanism for collecting tagged lamprey thus allowing an evaluation of the effectiveness of the antennae at detecting tagged lamprey. We will also collect untagged “wild” lamprey which can be tagged and used in subsequent trials and will provide information on the response of the “wild” spawning migration to the pheromone application. This provides another response variable and the ability to compare the behavior of our test animals.
Data analysis
Hypothesis 1-
πc= proportion of lamprey captured in Pond Brook without pheromone
πp= proportion of lamprey captured in Pond Brook with pheromone
H0: πc = πp
Ha: πp > πc
Response variableswill be; 1) the proportion of tagged sea lamprey that pass a detector array and the proportions of 2) tagged and 3) wild migratory lamprey trapped in each stream during treatment conditions (pheromone application) and control conditions (no pheromone application). ‘Successful’ outcomes will be a higherproportion of tagged and or untagged lamprey that move into Pond Brookversus Malletts Creekduring the “on” phase of each trial than during the “off” phase, and a higher capture of tagged and untagged lamprey in the Pond Brook trap above which pheromone is being applied relative to the trap in Malletts Creek without pheromone. Additional data of interest will be the rate of movement of tagged lamprey as determined by time it takes to pass an antenna. A biologically significant result would require 70 – 85% reduction in the numbers of lamprey migrating up Malletts Creek (Jones et al. 2003).
Proportional data from PIT tag receivers and trap catch will be analyzed using a χ² test to determine if the migratory lamprey distribute themselves according to the presence of pheromone.
Hypothesis 2-
τc= time to pass upstream antenna in Malletts without pheromone
τp=time to pass upstream antenna in Malletts with pheromone
H0: τc = τp
Ha: τp > τc
Rate of movement to the upstream antenna in Malletts Creek will be analyzed using a t-test to determine if there is a significant difference in the rate between on and off trials.
Hypothesis 3-
H0: πp < .70
Ha: πp ≥ .70
Since effect size is important, a 70% reduction in Malletts Creek is needed for control to be effective. A χ² test will be used to determine if the observed proportion of lamprey entering Pond Brook meets the criteria for a potential management application.
Hypothesis 4-
NT = Total number of lamprey captured
NH = Historical average of lamprey captured
H0: NT = NH
Ha: NT > NH
A t-test will be used to compare the total numbers of lamprey captured in Malletts Creek and Pond Brook to historical averages.
The number of tagged and untagged lamprey captured in traps will also allow for a mark-recapture estimate of the magnitude of the spawning run on Malletts Creek. This will give us an additional estimate of spawner abundance to determine if a biologically significant portion of the run is diverted into Pond Brook during the “ON” phases. We will also attempt to characterize the behavior of tagged lamprey, in terms of confusion or motivation, as indicated by PIT tag reads at antennas.
Management Implications
There are several possible applications for employing migratory pheromone in sea lamprey control. Sorensen and Vrieze (2003) identify five potential strategies; 1) trapping migrating sea lamprey in a lake, 2) trapping migrating sea lamprey in a river, 3) diversion into streams unsuitable for sea lamprey spawning or lavae?, 4) diversion within streams, and 5) disruption of migratory runs. This study is designed to address two of these scenarios (2 and 4) in part. First, pheromones could be used to activate streams where traps could be effectively used for collection and removal of migrating sea lamprey prior to spawning. The use of seasonal barriers and traps has been effective in controlling larval populations of sea lamprey on a limited basis (Bouffard et al. in prep). Second, if the distribution of migrating lamprey can be effectively manipulated within a tributary system using pheromones, migratory runs could be diverted into smaller tributaries of larger systems, where trapping and removal could be employed as a control technique. Alternately, redistribution of larval populations into different tributaries could provide considerable advantages for chemical control by eliminating large stretches of rivers which could be costly to treat with lampricides.
This will be the first experiment where migratory pheromones are applied to a side tributary of a stream which has a resident larval population and therefore a background odor. It is important to determine if this background odor can be overridden with the application of pheromones at higher concentrations. If successful, this study would be an important step in the development of control techniques using pheromones and would have considerable implications for the management of sea lamprey in Lake Champlain and in the Great Lakes.
Figure 1. Diagram of field site showing locations of antenna arrays and release location.
Figure 2. Diagram of flume for controlled environment experiments.
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