RAMP data from field observations

Data for fish and crabs show examples of how reflex impairment observations can be made during commercial fleet operations to estimate unobserved mortality.  With RAMP methods, experiments can be conducted by the fleet that identify important controlling factors for decreasing unobserved animal mortality from discarding and interactions with fishing gear. 

Davis 2002

Even though RAMP estimates for mortality in commercial fisheries may not presently have absolute accuracy, they are useful for identifying parts of fishing processes that have major impacts on unobserved mortality rates.  Carefully designed and fleet-inclusive observations of reflex impairment and RAMP during fishing, as well as experiments with gear configurations and deployment in different areas, seasons, and times of the day will certainly contribute to essential knowledge for fisheries management and allocation of scarce research and fishery resources.  Other sources for variability in RAMP scores and mortality estimates (predators, disease, and observer bias) may be evaluated using concurrent tagging of animals to compare RAMP estimated mortality rates versus tagging estimated mortality rates.  

Seasonal patterns were evident in RAMP scores and discard mortality for fish caught in the Georges Bank scallop fishery; for winter flounder,

and for yellowtail flounder.

Seasonal patterns of mortality related to windchill and temperature were evident in the snow crab fishery.

An experimental study by Rose et al. 2013 quantified RAMP and unobserved mortality for crabs that were encountered by bottom trawls in the Bering Sea. 

RAMP and corresponding mortality rates for snow crab, southern tanner crab, and red king crab.

Effects of trawl gear components on RAMP for crabs.

Effects of trawl gear components on mortality rates for crabs.

Examples of reflex impairment data

Here are some examples of the effects of stressors on reflex impairment and how individual reflex impairment can vary among fish species.

Atlantic cod reflex impairment and mortality occurring after exposure to air for 5 to 20 min Humborstad et al. 2009.

Coho salmon reflex impairment occurring after time in a landed seine net from 3 to 15 min Raby et al. 2012.

Proportion of coho salmon individual reflex impairment at various levels of RAMP Raby et al. 2012.

Proportion of walleye pollock, coho salmon, and northern rock sole individual reflex impairment after towing in a net for 5 min followed by exposure to air for 0 to 15 min. Pacific halibut were towed in a net for 240 min followed by exposure to air for 10 to 30 min Davis 2007.

Proportion of walleye pollock, coho salmon, northern rock sole, and Pacific halibut individual reflex impairment in order of impairment and contribution to total impairment Davis 2010

Different patterns of individual reflex impairment among species suggest hypothesis tests that could be made about the relative development and importance of individual reflexes and how these relate to life history patterns. Walleye pollock and coho salmon are pelagic species while northern rock sole and Pacific halibut are benthic species.

Science and reflex impairment testing using RAMP

RAMP is a method for testing whole animal reflex impairment and can be used in scientific inquiry.  The method of science includes three key elements for the explanation and prediction of observations through hypotheses.  Scientific hypotheses are 1) confirmable, meaning that they can make unexpected predictions about outcomes of experiments; 2) falsifiable, meaning that they can be tested by specific experiments; and 3) unique, meaning that they are the simplest and most plausible explanations for experimental observations.

Scientific hypotheses and explanations that fail these three tests are abandoned as not consistent with rational thought and discourse.  There are, of course, unlimited non-rational explanations for observations and perceptions and these are the purview of imagination, art, religion, philosophy, and politics as subjects of opinion and not necessarily open to testing, validation, and changing minds through discourse.  Lots of fun can obtain from imagination and humans consistently enjoy imaginative play outside of the realm of scientific inquiry.

The RAMP method certainly contains the three tests for scientific inquiry.  The RAMP reflex tests are specific observations of animal responses that are stable and repeatable under control conditions and that change as animal vitality is impaired.  These tests are designed as a quantitative language to communicate whole animal vitality states to observers.  RAMP can be used to confirm new predictions about animal vitality, morbidity and mortality.  Animals can be experimentally exposed to stressors (light, sound, temperature, hypoxia, injury, handling, and xenobiotics) in a wide variety of ecologically and economically relevant settings.  Specific stress outcomes (vitality, recovery, morbidity, and mortality) can be predicted using RAMP and then validated by observations through time and space.  RAMP can be used to falsify specific hypotheses concerning relationships among stressors, animal vitality, and stress outcomes.  RAMP can be used to identify unique explanations for experimental observations of animal outcomes in stressful conditions because of its singular expression of animal vitality, without being confounded with other factors, i.e., motivation, avoidance, attraction, size, sex, and species.

Additionally, RAMP is an interesting formulation of specific animal neurological, muscle, and organ reflex actions.  While RAMP accurately measures whole animal reflex impairment, the mechanistic details of reflex actions remain to be studied, as little is known about how stressors affect neurological function and associated reflex actions in different phyla.  The fact that reflex impairment is almost immediate after exposure to acute stressors suggests that neurological changes are important primary stress responses.  However, quantification of neurological dynamics is difficult and expensive because of their ephemeral nature.  Although rapid neurological changes in a stressed animal may be short-lived, RAMP data show that these changes exert profound control of reflex actions and later fitness outcomes.

Sorting wild-caught animals for high quality and live market

Many fishers are becoming aware of the value added to catch by controlling and accounting for animal vitality and food quality.  Fish and crabs that are stressed by capture and then transported to markets, either as live animals or freshly killed, have lesser value for discriminating consumers.  Vendors and educators for sea- and fresh-water food are developing programs to account for the methods and sustainability of food capture and handling.

Accounting for animal vitality and quality requires information about capture methods and associated impacts on ecosystem structure and function.  Sorting of catch is important for maintaining high market quality.  Rapid transport of catch to market can insure fresh product.  All of these aspects of fishing can benefit from the information that RAMP supplies.

Changes in the design of fishing gear and fishing methods that improve catch vitality and decrease bycatch can be tested and validated using RAMP to measure animal stress responses to the stressors of capture and handling.

Turtle escaping trawl

Fish or crabs can be captured and then transported to net pens for holding and later release onto markets.  Planned supplies of fresh product stabilize markets and increase the value of the catch.  These methods of capture and marketing require the availability of animals with best vitality and quality to reduce transport and holding costs associated with poor quality catch.  RAMP has been proposed for use in an Atlantic cod capture-based aquaculture system.

Capture-based tuna

Increasing use of combinations of wild fishing and aquaculture rearing requires information on animal quality, vitality, and fitness.  RAMP can supply this information used to test and validate the design and use of new aquatic food supply systems and sources.  Because RAMP is a cheap, easy, and immediate source of critical data on animal vitality and fitness, it can be an important component for efficient economies of operation in aquatic food production.

Capture-based aquaculture

The ornamental fish trade has a big impact on wild fish populations and is in critical need of information on fish vitality and fitness in capture, holding, transport, and marketing aspects of the industry.  RAMP can easily, effectively, and economically supply this needed information to improve the sustainability and ethics of this growing industry.

Ornamental fish trade

Handling, transporting, and processing animals often involves the use of anesthesia. Reflex impairment and RAMP can be used to assess the induction and recovery from anesthesia. 

Induction of anesthesia in fishes is described by loss of activity and responsiveness (Neiffer and Stamper 2009); “With proper dosing, induction with immersion drugs usually occurs within 5 to 10 minutes, but may take longer via other routes. Induction is marked by decreases in caudal fin strokes, swimming, respiratory rate, and reaction to stimuli; the drop in caudal fin stroke activity is usually the first sign, followed by loss of equilibrium and response to stimuli. At surgical anesthesia there is total loss of muscle tone and a further decrease in respiratory rate. A firm squeeze at the base of the tail can be an effective way to determine response to stimuli: if the animal does not respond, general anesthesia has taken effect (Harms 2003; Stetter 2001).”

Figure shows loss and recovery of swimming and equilibrium (behaviours), startle response to probe stimulation (responses), and vestibular-ocular response and rhythmic breathing (reflexes) associated with anesthesia induced in rainbow trout by MS-222 (Kestin et al. 2002). 

Concern about potential observer bias in measures for vitality

A paper by Benoît et al. 2010 summarized concerns about potential observer bias for scoring fish vitality.  The writers scored fish vitality using a "semi-quantitative measure based on four code levels of activity and injury":

A similar vitality scoring system with three levels of code; excellent, poor, and dead, is used to measure Pacific halibut bycatch viability in NE Pacific fisheries; see Appendices Q, R, S, T, U, and V in the 2012 Fisheries Observer Manual.  Manire et al. (2001) and Hueter et al. (2006) used a condition scoring system for sharks consisting of five levels of code; good, fair, poor, very poor, and moribund. Enever et al. 2009 used a condition scoring system for skates consisting of three levels of code; good, moderate, poor.

On their vitality method, Benoît et al. 2010 state:

"Because the scoring is based on a rapid visual and tactile assessment by observers, a large number of individual fish from a variety of fishing sets can be sampled. To the extent that observers obtain samples that are representative of the activities of a fishery, the observed frequency distribution of fish among vitality levels will reflect the overall survival potential of discarded fish, integrating over all of the various factors contributing to that survival. Furthermore, these studies typically yield sufficient sample sizes and contrasts in the factors affecting survival to allow for correlative analyses over the relevant ranges of a number of those factors. The principal disadvantage of these studies is the somewhat subjective nature of the scoring criteria, which could lead to differences between observers in their vitality assessments. A second disadvantage is the need for additional studies to relate vitality scores to eventual survival."

Their solution to potential observer bias is to use an appropriate statistical model for vitality:

"The study addresses the two disadvantages of vitality-score studies listed above. We propose the use of a mixed effects multinomial linear model based on proportional-odds (e.g., Agresti 2002, pp. 513–515), which is appropriate for modelling the ordinal vitality data and is a useful approach for addressing observer scoring subjectivity. We use this model to evaluate the effect of relevant factors believed to influence the survival potential (e.g., gear type, fish size, set duration, handling time) for eleven fish taxa. We also present analyses of an experimental study aimed at relating vitality codes to eventual survival."

From the perspective of RAMP, sources for potential observer bias are addressed by shifting measured variables from general observations of activity and injuries to observations of specific reflex actions (and injuries where appropriate). These specific reflex actions can be consistently quantified as present or absent and combined in a quantitative reflex impairment score, RAMP. Studies of potential observer bias for scoring reflex actions can be made by comparing RAMP scores among teams of observers in systems of interest.  

A second disadvantage for vitality scores is that they are not direct measures for animal fitness. This seeming deficit is a key need in all research on measures for vitality and stress.  Scores for animal activity, injury, physiology, or reflex actions can be used directly to compare the relative effects of stressors on stress induction. Ultimately these vitality scores and other measures of stress can and should be linked to fitness outcomes that cannot be observed directly, i.e., delayed mortality or recovery.  Tagging animals and tracking their fates in open or closed systems can supply information for linking vitality measures to fitness outcomes over a wide range of temporal and spatial scales in individuals and populations.

Using RAMP in fishing gear design

RAMP has been used in the Bering Sea to measure real-time reflex impairment and predicted mortality in tanner and snow crabs contacted by bottom trawl footropes that are part of trawls used to capture walleye pollock.  The RAMP data can be used in real time to adjust trawl performance in an effort to engineer trawls for reduced crab mortality.  Rapid access to reflex impairment data during the research cruise means that the researchers and fishers can adjust and test trawl performance immediately, without waiting for an additional future cruise.

Measuring crab size

Testing crab reflex action, also see here

Reflex impairment related to pollutant concentrations

Reflex impairment can be used to test for the behavioral and fitness outcomes of exposure to possible pollutants.  A recent study tested effects of triclosan on reflex impairment in an estuarine fish, Atlantic croaker:

"The effects of triclosan on reflex responses and anti-predator behavior in an estuarine fish

Tiffany L. Hedrick-Hopper and Sandra L. Diamond, Department of Biological Sciences, Texas Tech University, Lubbock, TX Background/Question/Methods Triclosan is a common antibacterial compound found in an increasing number of personal care products including toothpastes, deodorants, and soaps. Despite partial removal by wastewater treatment plants, an increasing amount of triclosan is entering watersheds where it can have significant effects on aquatic organisms. Even at low levels, triclosan negatively impacts thyroid homeostasis in anurans and fish, and it can decrease startle responses and activity levels in anurans. The purpose of this research was to investigate the effects of triclosan on reflex responses and anti-predator behavior in juvenile Atlantic croaker (Micropogonias undulatus), an estuarine fish. Sixty Atlantic croaker were held in individual tanks and randomly assigned to be fed a diet of either normal food pellets or pellets impregnated with 50 ppm triclosan for 14 days. Both prior to and immediately following the 14 day exposure, fish were tested for a suite of reflex action mortality predictors (RAMP) and were subjected to a video-recorded 30 second simulated predator attack. Videos were then analyzed for the specific strategies (run, hide, cut across tank, turn gambit) employed by the fish before and after exposure. Results/Conclusions We found that fish exposed to triclosan were significantly more likely than control fish to exhibit reflex impairment. Specifically fish lost the dorsal spine erection response, meaning that they did not raise their dorsal fin when the fin was flattened. Reflex impairment is correlated with increases in overall fish stress and mortality outcomes. Treated fish also experienced significant shifts in their anti-predator strategies. Triclosan-exposed fish spent significantly more time in their post-exposure test hiding from the simulated predator than fish in the control group. In some cases, fish continued to stay stationary even as the simulated predator touched them. The results of this study indicate that triclosan does impair fish reflexes and creates shifts in the strategies used by croaker to escape their predators. Since these schooling fish have been shown to exhibit dominance hierarchies, triclosan may affect social patterning in Atlantic croaker. These behavioral effects may have important implications not only for croaker and similar fish species but also for croaker predators such as bottlenose dolphins as contaminated fish may be easier prey, leading to increased predator body burdens." This study used free-swimming fish and video analysis of reflex responses.  The RAMP approach can easily be adapted to pollutant research and aquaculture settings for efficient, real-time monitoring of supply waters and sediments, as well as health conditions for animal rearing.  In the basic research context, neurobiological studies of zebrafish have made use of reflex testing in pharmacology and toxicology laboratory settings.

Sharks, Rays, and Chimaeras: reflex impairment and delayed mortality

Recent field research on sharks, rays, and chimaeras quantifying post-capture survival (PCS) has shown that:

 "As in any risk assessment, the methodology presented in this study is a first step to indentifying which species are more at risk. It provides an alternative and demonstrates that we are able to gain comparable knowledge on PCS for a large number of species from observations conducted on board commercial fishing vessels. All species showed little variation in the ‘wounds and bleeding’ and ‘sea lice’ indices values, suggesting that these indices could be omitted from future assessments under similar conditions. On the contrary, the ‘activity and stimuli’ index would be a cost-effective method for assessing the general condition of an animal in order to predict subsequent events in its life. For example, release condition (an index comparable to the activity and stimuli' index) was one of the best and most consistent predictors of the PCS of tropical reef fish and at the same time simple enough to be used by recreational fishers for a broad assessment of species."

The study makes concluding remarks: 

"A very large proportion of chondrichthyan global catches is discarded [5], [52] though little is known about the fate of discarded individuals. Hence, PCS information is rarely considered as part of the strategies addressing the management of discarded chondrichthyan species. Given that chondrichthyans remain a low priority for fishery management agencies in general, cost- and labour-intensive research on the broad range of species taken in commercial fisheries may not be conducted in the short term. However, the current change in natural resource management objectives from single-species to ecosystem-wide objectives warrants a multi-species assessment of PCS. Yet multi-species assessments are more difficult, and finding more cost-effective and priority driven methods is important because chondrichthyans continue to be depleted and time and funding for comprehensive data collection is limited [30]. Our study provided species-specific estimates of PCS, showing that these estimates varied among species, but they were generally high for most discarded species. The risk-assessment approach is simple and easy to implement in the onboard observer programs currently monitoring commercial fisheries around the globe, allowing the identification of species of conservation concern, and the prioritization and better direction of research and conservation effort.

Scoring reflex actions

Once we have learned how to "tickle" animals to stimulate a variety of reflex actions, we can consider how to score these actions.  Individual reflex actions can be scored either as present or absent, or can be scored according to their strength of response. We might think that scoring a reflex action by its strength would be the most useful, as it gives a continuous range of values from full strength in large powerful animals through weaker in healthy smaller animals, to weak in stressed large and small animals, and absent in fully impaired large or small animals. A disadvantage of this strength scoring approach is that both animal vitality and animal size can control the strength of reflex action. We are interested in an index for vitality, not confounded by animal size, so using strength of reflex action is not going to be appropriate.

To factor out the effects of animal size, we are left with scoring a reflex action as present or absent.  This may initially be confusing, with the question remaining, is the reflex present or not? I use the "rule of doubt" which says that the reflex is present if it is strong and clearly evident. It is scored absent if there is a question about its presence, whether it is weak or not clearly evident.

Scoring an individual reflex present or absent only gives us a qualitative measure and we are looking for a quantitative measure of vitality. We then move from the perspective of scoring individual reflex actions to a whole animal perspective, which includes many reflex actions, arising from combinations of various physiological, neural, organ, and muscle systems. If we measure the presence or absence of several reflex actions, representing combinations of various metabolic systems in an animal, then we can combine these scores into an integrated measure of reflex impairment that varies continuously and reflects the integrated nature of metabolic systems included in an animal. The combined scores for reflex impairment are then used to construct a RAMP curve which models the effects of stressors on animals through a range of stress and reflex impairment, as related to potential mortality.

An advantage of using RAMP is that the animal tells its story of stress-related impairment using systems that naturally integrate function as a whole animal. The animal communicates its vitality state directly through the language of reflex actions in response to appropriate stimuli, without the confounding effects of size and motivation.

RAMP: from intuition to science

Lets begin with fish, but the discussion applies to all other animals that have reflex actions.  Every fisher, commercial or recreational, intuitively knows and expresses opinions about the vitality of their fish, either in the water or caught. Excitedly proclaiming fish on and then proceeding to catch the fish, admire its size, and then release, sell, or eat the fish. These intuitive observations are grounded in our sense of vitality that is an expression of activity and responsiveness.

Intuitive notions are great for telling fish stories and are notoriously fallible when the size or fight of the fish in question is described to other bystanders. But these notions can lead to a quantitative expression of animal vitality that is grounded in solid, repeatable, and predictive science. How do we do this?

Vitality can be an expression of activity, which is diminished in stressed, lethargic fish.  Stress is an adaptive response to stressors. When fish are stressed too much or for too long, they can become diseased or die, states that do not support healthy populations and species diversity. So this loss of vitality that we intuitively observe can have profound consequences. To understand and ameliorate these consequences, we need good quantitative science.

For the purposes of describing and quantifying animal vitality and its inverse, mortality, we can start with animals in good condition and health that have a full suite of reflex actions and then study how stressors impair reflex actions until the end point of death. We use the presence or absence of reflex actions because these are fixed involuntary actions that are directly related to vitality and not subject to the effects of animal size and voluntary, complex behaviors such as feeding, social interactions, predator-prey interactions, migration, and sex, which can be modified by temperature, light, food availability, motivation, avoidance, and attraction.

We use a calculated quantitative index of reflex impairment, RAMP, that combines the presence-absence scoring of several reflex actions. RAMP is an integrative index that communicates the vitality of a whole animal. Similar reflex-based indices are used in human medicine to evaluate general health, neurological condition, and potential outcomes for coma and other non-communicating patients, as well as for triage of emergency patients.

Identifying appropriate reflex actions is where the imagination expands. We have got to figure out how to "tickle" the animal. What stimuli make it respond in the fixed involuntary patterns we call reflexes? Appropriate stimuli and testing modalities depend on the size of the animal and the logistical constraints of the situation. There are many human examples for inspiration.

Lets look at reflex actions through a continuum of animal size and activity for examples.  This list is by no means complete. Reflex actions can be tested in fish larvae by observing free swimming animal startle, orientation, and avoidance in response to light, sound, food scent, and touching with a probe.  For juveniles, fish can be restrained and tested for body flex upon restraint where fish attempt to escape when restrained, dorsal fin erection in which the fins become erect when fish are restrained, operculum and mouth closure where the operculum or mouth clamps shut when lifted or opened, the gag response where the fish opens its mouth and flexes the body when the throat is stimulated and the vestibular–ocular response (VOR) shown by eye rolls when the body is rotated around the long axis. For free swimming fish, studied reflexes included orientation where the fish should normally be upright, righting reflex where the fish returns to an upright position and the startle response in which the fish shows rapid forward motion in response to stimuli. Adult fish can present special problems because of their strength and other approaches for free swimming fish are described in another post. Sharks and other dangerous toothy or spiny animals especially need imaginative approaches to testing reflex actions.

Once a suite of reflex actions can be consistently observed and easily quantified, then building a RAMP curve can be accomplished and quantification of reflex impairment, vitality, and prediction of mortality is made possible. The RAMP method and curves developed then allow for the systematic investigation of the effects of stressors and stress in animals and systems of chosen interest. RAMP results can be compared and contrasted with concurrent results from physiological and physical injury studies in an effort to synthesize multivariate solutions to a continuum of important basic and applied questions. These questions may include understanding reflex biology, stress biology, fisheries management, bycatch reduction, animal health, population dynamics, aquaculture practices, migration biology, reproductive biology, and conservation biology to mention a few. 

RAMP validation

Initial research with RAMP was designed to show proof of principle for the efficacy of using reflex impairment to measure stress induction and predict mortality in fish and invertebrates.  These results are reviewed by Davis 2010 and Stoner 2012.

A next step in RAMP research is to validate the method under field conditions related to fitness outcomes such as migration and survival.  Examples of field validation are beginning to appear for coho salmon in Raby et al. 2012 and dungeness crab in Yochum 2012.  Validation studies use a suite of experimental methods to construct RAMP curves, capture experimental subjects, tag and release the captured animals, track their movements, behavior, and survival, and correlate RAMP scores with delayed mortality in field settings.  Of course, when RAMP is used in closed systems like aquaculture, live capture, and smaller lakes, validation is simpler since animals are retained and can easily be monitored and tested throughout their life.

Additional validation studies can also be conducted to document the possible effects of capture or rearing conditions, ontogeny, injury, and predator abundance on RAMP curves for different species.  In general a RAMP curve should be constructed using animals exposed to the types and intensities of stressors under which the RAMP curve will be used.

Research on the mechanisms of reflex impairment induced by stressors can be undertaken which will contribute significantly to understanding the integration of neural, muscular, and organ systems in animals.  The study of reflex impairment contributes a different and valuable perspective to standard models for stress, disease, morbidity, and death in animals.  While significant work has contributed to understanding reflex actions and their relation to health in humans,  impairment of animal reflex actions and their fitness outcomes are less studied.

Anatomy of a RAMP curve

RAMP curves are different for each animal species. These differences are probably related to differing abilities for adaption to stress. Within a species, the RAMP curve is similar or identical among different  animal sizes and stages of ontogeny, assuming that the component reflex responses are developed.  In RAMP, individual reflex actions are scored as present or absent and then combined to express the proportion of reflex actions impaired. The presence-absence scoring factors out the effects of animal size and development. A different way to score would be to measure strength of reflex actions, but this would vary with animal size and would not independently measure vitality. Other ways of scoring for RAMP, which could include injury, will be discussed in a separate post.

RAMP data can be fitted to logistic curves to express the relationship between reflex impairment and delayed mortality.

Restrained Atlantic cod, Humborstad et al. 2009

 Restrained fish, Davis 2009

 Unrestrained fish, Davis 2009

Spot prawn, Stoner 2012

The RAMP curve begins with a control condition of no reflex impairment and no mortality. The left hand curve tail continues at no mortality as reflex impairment increase, i.e., the animal becomes less responsive and more lethargic, in a zone of sublethal stress effects. At the first inflection point, as reflex impairment increase, probability for mortality appears and rapidly increases with small increases in reflex impairment, in a mixed zone of sublethal stress effects and morbidity. Finally, a second inflection point is reached at the right hand curve tail and high probability for mortality is evident at high levels of reflex impairment. The RAMP curves vary among species while maintaining these logistic characteristics.

Tanner crabs and snow crabs, Stoner 2008

A study with crabs, in which injury was also scored, showed a family of RAMP curves that varied over injury intensity for Tanner crabs, but not for snow crabs. For Tanner crabs it is useful to include injury scoring in the RAMP calculations, as reflected in the surface response curve shown.

Uses for RAMP

RAMP measures reflex impairment and predicts delayed mortality in animals. The method is a real time, cheap, effective, and easy way to monitor animal vitality, stress, disease, morbidity, and delayed mortality.  How is this useful?

In fisheries, non-target species of fish, invertebrates, birds, amphibians, reptiles, and mammals (bycatch) are captured and released for a variety of reasons, usually related to management requirements or economic factors.  The release of these animals can result in large, unquantified amounts of fishing mortality that causes uncertainty in fisheries management and wastes valuable lives and resources.  RAMP can give immediate real time data on the vitality and potential delayed mortality of captured animals.  This data can be used in real time to evaluate and adjust fishing practices to improve bycatch survival and to quantify bycatch fishing mortality.

In live fisheries, animals are captured and transported to net pens or land markets for food consumption or aquarium trade.  Sorting of animals for maximum survival can be quickly accomplished using RAMP.  This sorting saves valuable holding space, decreases shipping costs, and increases the value of catch by including only top quality animals.  RAMP also aids in perfecting capture practices that maximize animal vitality and survival.

In aquaculture, RAMP is useful for real time monitoring of animal vitality, stress, disease, and potential for morbidity and mortality.  Aquaculture depends on maintaining the highest health conditions for animals, as disease can be a major impediment to aquaculture ethics and economics.  RAMP monitoring can be automated in aquaculture settings by using computer aided reflex testing.  Stimuli such as flashing bright light, sound, and food scent can be administered randomly into holding areas and the reflex responses of free-swimming animals can be observed and analyzed by video systems.  Detection of impaired reflex responses can trigger alarms systems and bring personnel to adjust conditions for improved animal health.

In pollution research and monitoring, RAMP is a sensitive measure of animal vitality, stress, sublethal, and lethal effects of pollutants. A large body of research exists documenting effects of pollutants on volitional feeding, social behavior, and predator prey interactions. RAMP can be an even more powerful measure of pollution effects than volitional behavior, since reflex impairment is directly related to animal vitality, without the modifying effects of motivation, size, and sex.

Examples of RAMP research

Examples of RAMP are beginning to appear in the scientific literature and on the internet.

Here are links to some of these studies in fish - Atlantic cod, Humborstad et al. 2009; rockfish, Hochhalter 2012; yellowtail flounder, Barkley and Cadrin 2012; red snapper, Campbell et al. 2009, Campbell et al. 2010, 2010; coho salmon, Raby et al. 2012; review, Davis 2010.

Here are links to some of these studies for invertebrates -  tanner and snow crab, Stoner et al. 2008, Stoner 2009; review of crustaceans, Stoner 2012; spot prawn, Stoner 2012; dungeness crab, Yochum 2012.

Here are some RAMP curves from studies.

Stoner 2008

Stoner 2012

Davis and Ottmar 2006

Davis et al. 2009

Why does RAMP work?

RAMP is a whole animal quantitative measure of health and vitality. It integrates several reflex actions that are combinations of neural and muscle function which are immediately responsive to the effects of stressors.  When an animal is exposed to stressors and becomes stressed, various physiological, organ, and behavioral systems respond in adaptive ways to compensate for the disturbance of stress. Initially these stress responses are beneficial, helping the animal avoid stressful situations and stimulating metabolism to support these adaptations. However if stress is prolonged, the animal begins to exhibit metabolic deficits and its health and vitality degrade.

An animal with disturbed states and degraded vitality can quickly become sick, moribund, and eventually die if stress persists at high enough levels. Prediction of animal death or recovery from stress requires measuring whole animal stress disturbances. Measuring disturbances of separate systems that make up the animal does not predict vitality and mortality because the whole animal is what dies, not the separate systems.

RAMP is a combination of several reflex actions that is an ideal predictor of whole animal vitality and mortality because it integrates the immediate effects of stress for the whole animal into involuntary fixed patterns of response that vary only with the vitality of the animal. If voluntary behavior is used as a predictor, other factors not related to animal vitality can control responses, making prediction of vitality difficult.  For example feeding and other social interactions can be controlled by motivation, resource availability, avoidance, and attraction. If component metabolic and organ systems are used as a measure, these do not reflect the whole animal vitality state because they exhibit peak responses to stressor intensities that are not related to stress levels in the whole animal.

Getting started with RAMP

How do you get started with RAMP?  Recently I sent an e-mail to a researcher with this question on bycatch in a purse seine.  My answer follows.  Good to hear from you.  Sounds like you have some bigger fish to work with.  Sample sizes depend on what you are going to accomplish and the effect size anticipated.  1) The first tasks are to get fish in good condition, lean how to hold them in good condition, and then to establish a RAMP curve.  Get as many fish as you can comfortably hold and maintain.  Then 2) establish reflex actions that can be tested consistently and that respond consistently and strongly.  Find as many testable reflex actions as you can as this makes a more refined RAMP score.  See Davis 2010 and Raby et al. 2012 papers for detailed information.  For larger salmon that are difficult to handle, Raby tested without a restraining device: 
"Each reflex was assessed categorically (0 = unimpaired, 1 = impaired) in a conservative matter – that is, if the handler had doubt as to whether the reflex was present, it was recorded as being impaired. Reflexes tested were the following: tail grab, body flex, head complex, vestibular-ocular response (VOR) and orientation. Presence of the tail grab response was assessed by the handler attempting to grab the tail of the fish with the fish submerged in water (in a fish bag or holding trough); a positive response was characterized by the fish attempting to burst-swim immediately upon contact. The body flex response was tested by holding the fish out of water using two hands wrapped around the middle of the body. The fish actively attempting to struggle free was characterized as a positive response. Head complex was noted as positive if, when held out of water, the fish exhibited a regular pattern of ventilation (for 5 s) observable by watching the opening and closing of the lower jaw. VOR was observed by turning the fish on its side (i.e. on a lengthwise axis) out of water. Positive VOR was characterized by the fish’s eye rolling to maintain level pitch, tracking the handler. Finally, upon release, each fish was placed upside-down in the river just below the surface: a positive orientation reflex was noted if the fish righted itself within 3 s. The entire reflex assessment took 20 s to complete and was always conducted on fish upon release. If a fish was too vigorous to allow researcher handling and assessment of reflexes, it was assigned an unimpaired status for all reflexes."  
Then 3) once consistent reflexes are identified and tests are finalized, a RAMP curve is constructed which establishes the relationship between reflex impairment and mortality.  This is the scoping process that establishes what stressor intensities will kill fish and what are sub-lethal and produce lesser RAMP scores.  This process involves exposing fish to a range of capture stressor intensities of interest.  For purse seine capture this may include net abrasion time (extent of injury) and air exposure time (extent of hypoxia) and temperature if there are enough fish to test.  The stressor range should induce reflex impairment and mortality that ranges from no impairment and mortality to complete impairment and mortality.  In this way, the RAMP curve covers all possible outcomes for the fish.  If possible, the most efficient use of limited fish is to expose and tag individual fish so that for each individual, a RAMP score can be assigned to a specific stressor intensity.  4) Once you have a RAMP curve, you can test fish in the field for RAMP, note their capture conditions, and predict mortality based on RAMP and also on capture conditions.  

Initially you will need at least 10 fish to construct a RAMP curve, more (20 fish) is better as it will give you a curve with tighter confidence bounds.  For a simple example lets say that fish die after 15 min in air.  So exposures might look like this: fish 1- 0 min air, fish 2 - 2 min air, fish 3 - 4 min air, fish 4 - 6 min air, fish 5 - 8 min air, fish 6 - 10 min air, fish 7 - 12 min air, fish 8 - 14 min air, fish 9 - 16 min air, fish 10 - 18 min air.  If injury is important you might do a combination of net holding and air exposure.  Expose the fish to the assigned stressor, test the fish for reflex impairment, then hold the fish and observe for delayed mortality.  Holding time vary with species and usually range from 3-11 days.

The beginning of RAMP

I was researching the effects of capture and release of non-target commercially important fish species, commonly know as bycatch. The research was aimed at understanding what happens to fish when they are captured in commercial fishing gear and then released because of fishing regulations. The question is important because fish that are stressed by capture and release often show delayed mortality and this source of mortality is difficult to measure and account for in management of fish stocks.  Results of this research were summarized in an article published in 2002.

Because delayed mortality cannot be observed directly, some kind of measure of the animal's vitality prior to release needs to be developed for prediction of delayed mortality. Our team tried to relate blood plasma variables, measures of physical injury, and complex voluntary behavior with delayed mortality. However these measures were not correlated with immediate or delayed mortality.  

One day while watching fish recover after exposure to simulated fishing capture, I began to think about the fact that the intensity of fish involuntary behavior after capture appeared to be negatively related to the intensity of the capture stressor. Fish subjected to more intense capture stressors became more lethargic and their involuntary behavior was more impaired. These involuntary activities are known as reflex actions and represent fixed involuntary actions that occur in response to external stimuli such as gravity, bright light, loud sound, or touch. The reflexes can be part of important behavior such as startle, orientation, predator avoidance, feeding, migration, and sex.

So I decided to measure and quantify reflex actions and to try to correlate impairment of these actions with delayed mortality. This idea of measuring a suite of reflex actions and correlating the measure with mortality was very successful and I called it RAMP - reflex action mortality predictor. The development and deployment of RAMP is described here, here, and here.

Since that original work, other researchers have used the RAMP approach to quantify stress and predict delayed mortality in a variety of fish and invertebrate species. I will describe their work in future posts.