Belly up: Righting reflex action time to recovery correlated with delayed mortality?

Upside down fish in market tank (Hong Kong)

RAMP incorporates presence/absence of several reflex actions and injuries to measure vitality impairment and potential delayed mortality. A simpler method may be possible by measuring time for recovery of orientation when fish are placed upside down in water. This method can be tested.

Place a fish upside down in water and observe the time until the fish returns to normal orientation. This duration is a measure of vitality impairment. Longer recovery times indicate greater vitality impairment and data can be included in statistical models for relationships among fishery stressors, injury, righting time, and delayed mortality. We can test the relationship between righting impairment and delayed mortality. 

Righting reflex action is a central behavior that is the nexus of neural, muscle, and organ actions and is intimately linked with loss of physiological regulation associated with stressor exposure.  Olfactory impairment is another example of a central function that is correlated with delayed mortality (in humans, Pinto 2014).

Body orientation is a sensitive measure of fish consciousness. Presence or absence of righting can be included in the RAMP score. Loss and recovery of orientation is a well known symptom for induction of and recovery from fish anesthesia and is used as an indicator of morbidity and delayed mortality in stress experiments (Davis and Ottmar 2006, Szekeres et al. 2014, Raby et al. 2015).  

Measuring replicate animals for the time to righting recovery and delayed mortality after a stressor experiment can test the correlation between righting impairment and delayed mortality. If the correlation between righting and delayed mortality is valid and strong, then we have a rapid method for predicting discard mortality on board fishing vessels without need for holding or tagging fish to confirm their survival. Research groups on fishing vessels can observe fish during catching, landing, sorting, and discarding under differing stressors; seasons, water temperatures, tow durations, catch quantities, species mixes, and sorting times.

Measuring and scoring vitality impairment

Associations between vitality impairment and mortality in Tanner and snow crab; Stoner et al. 2008.

At the risk of repetition and irrelevance, I will repeat my short history with vitality impairment and mortality.  I began by trying to find out what kills fish. The word fish applies to all animal types in a fishery. We chose to do this in the lab for control reasons, given the common confounding of stressors in fisheries (Davis 2002). All sorts of objections were made by field people that the work was irrelevant because it did not include field conditions. Well we focused on the fish and their capabilities, in an effort to formulate hypotheses that could be tested in the field. We found that each species and size of fish has different sensitivities to stressors and that stressors of importance were different for species.  We also found that some stressors (temperature and hypoxia) could kill fish without apparent macro-injury. After killing many fish, we endeavored to identify characteristics (traits) of fish and fisheries that could be correlated with mortality as predictors, given the difficulty of holding fish and measuring delayed mortality in the field. We tried many traits; muscle and plasma physiology, stressors, volitional behavior, injury, and reflex actions.

Effects of fishing gear, temperature, and fish size on sablefish mortality, NOAA.

Muscle and plasma physiology were not correlated with mortality because these are alarm responses that can be adaptive or maladaptive responses to stressors. In specific contexts, lactate and CO2 may be useful where hypoxia or fatigue are a concern.  

Stressors are an approach that has garnered enthusiasm. However their effects can be confounded and difficult to model given the relatively unlimited combinations of factors that are possible in a fishery; seasonal effects, gear type and deployment times, catch type and amount, sorting and discarding.  

Volitional behavior is not correlated with mortality because it is subject to variation that is not directly linked to mortality, such as changes in perception, motivation, fear, and attraction; all which confound the relationship of behavior and mortality.  

Injury is often correlated with mortality, especially in accidental death.  However not all mortality is correlated with injury, as in the effects of temperature and hypoxia, for which micro-injury may be evident (apoptosis) but difficult to measure in the field. Often the effects of injury, temperature, and hypoxia are confounded making interpretations of their effects on mortality difficult.

Single reflex actions are often not correlated with mortality when they are part of systems not central to body function and regulation directly related to mortality. They may be important for complex behaviors; predator avoidance, feeding, habitat choice, migration and these can have indirect effects on mortality. 

We reasoned that fish had a quality called vitality and that vitality was correlated with mortality. However, for an individual fish this relationship is binomial. The fish is alive or dead. So decreasing vitality results in sublethal effects on behavior until a threshold is reached and the probability for death increases rapidly. In statistical groups of fish, decreasing vitality is log-linearly related to mortality.

At this point we chose to not measure the strength of reflex actions (because of confounded size effects); instead to score the presence or absence of several reflex actions as an expression of reflex action impairment and loss of vitality. Reflex actions are fixed behavior patterns based on neural, muscle, and organ functions which do not vary with changes in perception, motivation, fear, and attraction.  We chose to focus on several types of reflex actions to increase the probability that reflex actions key to body regulation were included. Later work has shown that the orientation reflex is such a key reflex, often correlated with morbidity and mortality associated with hypoxia and fatigue. 

Previous work with vitality scoring in fisheries had developed the semi-quantitative analysis method (SQA) of scoring fish activity and injury, which was used in tagging studies and in Pacific halibut discard mortality estimation (ICES 2014). The method observes the sequential loss of operculum clamping and startle to touch and increased injuries from minor to major and bleeding; with ordinal scoring (1-4) for severity of impairment. The vitality score is readily incorporated in multivariate models that may identify stressors of importance to mortality and model mortality based on those stressors. RAMP can be scored in a similar manner as SQA and included in multivariate models. RAMP scores severity of impairment by noting the sequential loss of several types of reflex action and inclusion of injury types. Scores range from 0 to a maximum which is the number of trait types observed for presence/absence. Strength of action and extent of injury are not included because of the confounding effects of size. The effect of size is included in the model explicitly as fish length or weight. Smaller fish will have more vitality impairment than larger fish, when exposed to equivalent stressors.

RAMP is a simple extension of the SQA concept that includes more testing for reflex actions. SQA and RAMP are similar scoring systems that differ by emphasis. SQA and RAMP score activity, responsiveness, and injury to quantify levels of vitality impairment. RAMP simply includes more information about types of reflex actions in an effort to include reflex actions that are central to body regulation over a range of stressor conditions.

The primary reason for inclusion of more reflex action information in RAMP is the observation that some reflex actions are central expressions for status of body regulation. Given the binomial nature of mortality observations, we need to know why fish die. They die for many reasons which all seem to point to the loss of physiological regulation; either homeostatic or allostatic regulation. How do we measure regulation?  Allostasis shows us that consideration of homeostatic set points is not sufficient to predict mortality. My view is that vitality is correlated with physiological regulation and that impairment of vitality and regulation leads to mortality when physiological bounds of the species are exceeded. Until we can directly measure the causes for mortality, we rely on measures for vitality based on activity, responsiveness, and injury.

For predicting mortality, I chose to measure vitality over modeling stressors because of the direct relationship between vitality impairment and mortality. Stressor interactions in fisheries can make interpretation of stressor effects on mortality difficult to interpret. Information about stressors in multivariate models for mortality can be used to identify changes in the design of fishing gears that reduce bycatch mortality. Then vitality impairment can be used to evaluate reduction in discard mortality associated with new gears.

Survival of schooling small pelagic fish discarded from purse seine fisheries

Greenback horse mackerel, Trachurus declivis 

Purse seine fishing

Vitality impairment and RAMP can be used to determine survival of discarded schooling small pelagic fish in purse seine fisheries.  When catch is too large, fish are “slipped” for the net and discarded. These discarded fish are usually exposed to some level of hypoxic conditions associated with crowding in the purse seine. Elevated temperature in surface water may be a stressor. Skin abrasion and scale loss can occur in the net. Many small pelagic species (mackerel, sardine, anchovy, smelt, herring) caught in purse seines are obligate or facultative schoolers that reflexively form groups mediated by the optomotor response. Vitality impairment can be tested for individual fish or groups. See Davis and Ottmar, 2006 for testing groups of free-swimming fish. Schooling fish seek the company of species mates, so testing groups of schooling fish is probably the most informative method. How is this testing done and linked with delayed mortality in captive observation tanks?    

RAMP links vitality impairment scores with delayed mortality scores. The RAMP estimate for delayed mortality is only as good as the mortality estimates from captive observation or tagging experiments. How many replicates are needed in captive observation experiments? The purse seine schooling species need to be held in groups. A replicate group size of ten fish is good for schooling. These fish must be held in good water quality and circulation, in a circular tank size that allows schooling. For initial RAMP formulation, you will need 10 replicate groups of ten fish each. Observations of reflex action impairment and delayed mortality should be made over a range of stressor intensities that result in delayed mortality of 0 to 100%. Then replicate vitality impairment scores are linked with replicate delayed mortality scores to form the RAMP which can be validated with further experiments and replication.

Suuronen, 2005  Stressors in capture and escape of fisheries.

Fish can be sampled from any point on the fishing process, depending on the stressors of interest. Reflex action testing can be made on a group of fish held in a circular observation tank big enough for schooling (See Davis and Ottmar, 2006).  Possible reflex actions for testing include: orientation; schooling; rheotaxis; startle response to sound or light; swimming to bottom of tank. Injuries can also be noted; abrasion, scale loss. After testing the replicate group can then be placed in a holding tank and monitored for delayed mortality through five to ten days. 

Herring lose schooling, orientation, and tail beat frequency increases as the purse seine is drawn smaller (Morgan, 2014). Fatigue and hypoxia are possible stressors in purse seines (Tenningen 2014).

For discard species caught in purse seines that are not schooling fish, or are larger schooling fish, individual fish can be tested for vitality impairment.  Reflex actions tested can include: body flex, orientation, eye roll, operculum or mouth clamp, tail grab, righting, startle.  These fish can be tagged for identification and held together for five to ten days in tanks to determine delayed mortality.

Fisheries studies that have used and documented reflex actions or injury for vitality impairment testing

The table presents selected examples of reflex action & injury vitality impairment testing.

See also a list of reflex actions and injury that may be scored for vitality impairment.

Citations:

AFSC, Alaska Fisheries Science Center 2014 Observer Sampling Manual.

Fisheries Monitoring and Analysis Division, North Pacific Groundfish Observer Program. Seattle, WA.

Barkley, A.S. & Cadrin, S.X. 2012. Discard mortality estimation of yellowtail flounder using reflex action mortality predictors. Trans. Am. Fish. Soc. 141:638-644.

Benoît, H.P., Hurlbut, T., and Chassé, J. 2010. Assessing the factors influencing discard mortality of demersal fishes using a semi-quantitative indicator of survival potential. Fish. Res. 106:436-447.

Braccini, M., Van Rijn, J., and Frick, L. 2012. High post-capture survival for sharks, rays and chimaeras discarded in the main shark fishery of Australia? PLoS ONE 7: e32547.

Brownscombe, J.W., Thiem, J.D., Hatry, C., Cull, F. Haak, C.R., Danylchuk, A.J., and Cooke, S.J. 2013. Recovery bags reduce post-release impairments in locomotory activity and behavior of bonefish (Albula spp.) following exposure to angling-related stressors. J. Expt. Mar. Biol. Ecol. 440:207-215.

Brownscombe, J.W., Nowell, L., Samson, E., Danylchuk, A.J., & Cooke, S.J. 2014. Fishing-related stressors inhibit refuge-seeking behavior in released subadult Great barracuda. Trans. Am. Fish. Soc. 143:613-617.

Campbell, M.D., Patino, R., Tolan, J., Strauss, R., and Diamond, S.L. 2010.

Sub-lethal effects of catch-and-release fishing: measuring capture stress, fish impairment, and predation risk using a condition index. ICES J. Mar. Sci. 67:513-521. 

Campbell, M.D., Tolan, J., Strauss, R., and Diamond, S.L. 2010. Relating angling-dependent fish impairment to immediate release mortality of red snapper (Lutjanus campechanus). Fish. Res. 106:64-70.

Danylchuk, A.J., Suski, C.D., Mandelman, J.W., Murchie, J.W., Haak, Brooks, A.M.L., and Cooke, S.J. 2014. Hooking injury, physiological status and short-term mortality of juvenile lemon sharks (Negaprion bevirostris) following catch-and-release recreational angling. Cons. Physiol. 2:cot036.

Davis, M. W. 2007. Simulated fishing experiments for predicting delayed mortality rates using reflex impairment in restrained fish. ICES J. Mar. Sci. 64:1535-1542.

Davis, M.W. & Ottmar, M.L. 2006. Wounding and reflex impairment may be predictors for mortality in discarded or escaped fish. Fish. Res. 82:1-6.

Depestele, J., Buyvoets, E., Calebout, P., Desender, M., Goossens, J., Lagast, E., Vuylsteke, D., and Vanden Berghe, C. 2014. Calibration tests for estimating reflex action mortality predictor for sole (Solea solea) and plaice (Pleuronectes platessa): preliminary results. ILVO-communication. Report nr. 158. 30p. 

Diamond, S.L. & Campbell, M.D. 2009. Linking “sink or swim” indicators to delayed mortality in red snapper by using a condition index. Mar. Coast. Fish.: Dynamics, Manag. Eco. Sci. 1:107-120.

Donaldson, M.R., Hinch, S.G., Raby, G.D., Patterson, D.A., Farrell, A.P., and Cooke, S.J. 2012. Population-specific consequences of fisheries-related stressors on adult sockeye salmon. Physiol. Biochem. Zool. 85:729-739.

Hammond, C.F., Conquest, L.L., and Rose, C.S. 2013. Using reflex action mortality predictors (RAMP) to evaluate if trawl gear modifications reduce the unobserved mortality of Tanner crab (Chionoecetes bairdi) and snow crab (C. opilio). ICES J. Mar. Sci. 70:1308-1318.

Hannah, R.W. and Matteson, K.M. 2007. Behavior of nine species of Pacific rockfish after hook-and-line capture, recompression, and release. Trans. Amer. Fish. Soc. 136:24-33.

Humborstad, O-B., Davis, M.W., and Løkkeborg, S. 2009. Reflex impairment as a measure of vitality and survival potential of Atlantic cod (Gadus morhua). Fish. Bull. 107:395-402. 

McArley, T.J. & Herbert, N.A. 2014. Mortality, physiological stress and reflex impairment in sub-legal Pagrua auratus exposed to simulated angling.  J. Expt. Mar. Biol. Ecol. 461:61-72.

Raby, G.D., Donaldson, M.R., Hinch, S.G., Patterson, D.A., Lotto, A.G., Robichaud, D., English, K.K., Willmore, W.G., Farrell, A.P., Davis, M.W., and Cooke, S.J. 2012. Validation of reflex indicators for measuring vitality and predicting the delayed mortality of wild coho salmon bycatch released from fishing gears. J. Appl. Ecol. 49:90-98.

Stoner, A.W. 2012. Assessing stress and predicting mortality in economically significant crustaceans. Rev. Fish. Sci. 20:111-135.

Stoner, A.W., Rose, C.S., Munk, J.E., Hammond, C.F., and Davis, M.W. 2008. An assessment of discard mortality for two Alaskan crab species, Tanner crab (Chionoecetes bairdi) and snow crab (C. opilio), based on reflex impairment. Fish. Bull. 106:337-347.

Szekeres, P., Brownscombe, J.W., Cull, F., Danylchuk, A.J., Shultz, A.D., Suski, C.D., Murchie, K.J., and Cooke, S.J. 2014. Physiological and behavioural consequences of cold shock on bonefish (Albula vulpes) in The Bahamas. J. Expt. Mar. Biol. Ecol. 459:1-7.

Trumble, R.J., Kaimmer, S.M., and Williams, G.H. 2000. Estimation of discard mortality rates for Pacific halibut bycatch in groundfish longline fisheries. N. Amer. J. Fish. Manag. 20:931-939.

Yochum, N., Rose, C.S., and Hammond, C.F. 2015. Evaluating the flexibility of a reflex action mortality predictor to determine bycatch mortality rates: A case study of Tanner crab (Chionoecetes bairdi) bycaught in Alaska bottom trawls. Fish. Res. 161:226-234.

RAMP is a component of an integrated conservation approach to coho salmon bycatch mortality management

Raby et al. 2014

Results of Raby et al. 2014 demonstrate the integration of vitality impairment and coho bycatch mortality estimation and management.

“We have provided an estimate of bycatch mortality for an endangered population of coho salmon captured in an aboriginal beach seine fishery, based on three years of tracking fish released from the fishery.” 

“Among all the variables we tested as predictors of mortality, none were significant except for RAMP score, whereby fish with higher RAMP scores (more impaired) were less likely to be successful migrants (Table 3, Fig. 4).”

Distinguishing between natural mortality and bycatch mortality. 

“An alternate approach to calculating a bycatch mortality rate that attempts to distinguish bycatch from natural mortality, is to use RAMP scores and their mortality rates at each level of impairment, and assume negligible bycatch mortality for the fish that were least impacted (vigorous at release).”

“Since some in-river mortality is natural, there is a need to attempt to differentiate mortality caused by the capture itself. To do so, RAMP scores can be used whereby coho salmon released with little or no reflex impairment (vigorous) are assumed to experience no post-release bycatch mortality. Using that conservative assumption, the post-release mortality rate for those fish can then be used as a baseline within the data set. Additional mortality above that baseline that occurs at higher levels of reflex impairment can then be assigned to the fishery (see Fig. 4).”

Using RAMP to monitor condition of bycatch and improve their survival

“The expanded validation of the RAMP approach in the present study provides confirmation that this simple technique is ready for use in this fishery if needed (Raby et al. 2012). The observers in the fishery could easily be taught how to conduct RAMP assessments to monitor the condition of bycatch in real time, provide advice to their crews on how to improve fish condition, and make decisions about whether individual fish should be revived using recovery bags.”

Human delayed mortality can be predicted using olfactory impairment

Olfactory impairment in humans was measured by error rate in olfaction tests. Increasing number of errors in olfaction tests were related to increasing 5-year mortality rates in a logistic regression (PLoS ONE). 

The human logistic relationship between olfactory impairment and 5-year delayed mortality is a powerful method for predicting delayed mortality and is similar to other animal RAMP relationships between reflex impairment, injury, and delayed mortality. Olfactory impairment can be easily measured in human and animal clinical settings and can easily and automatically be measured in aquaculture contexts by analysis of animal distributions and activity in rearing facilities. Given the fundamental nature of olfaction, one would expect the relationship between olfactory impairment and delayed mortality to be generally present among animal phyla and this can be tested in clinical and field settings.

Pinto et al. 2014 state, “We are the first to show that olfactory dysfunction is a strong predictor of 5-year mortality in a nationally representative sample of older adults. Olfactory dysfunction was an independent risk factor for death, stronger than several common causes of death, such as heart failure, lung disease and cancer, indicating that this evolutionarily ancient special sense may signal a key mechanism that affects human longevity. This effect is large enough to identify those at a higher risk of death even after taking account of other factors, yielding a 2.4 fold increase in the average probability of death among those already at high risk (Figure 3B). Even among those near the median risk, anosmia increases the average probability of death from 0.09 (for normal smellers) to 0.25. Thus, from a clinical point of view, assessment of olfactory function would enhance existing tools and strategies to identify those patients at high risk of mortality.”

(PLoS ONE)

The human study controlled for the mortality effects of age, gender, socioeconomic status, and race. Additionally, “We excluded several possibilities that might have explained these striking results. Adjusting for nutrition had little impact on the relationship between olfactory dysfunction and death. Similarly, accounting for cognition and neurodegenerative disease and frailty also failed to mediate the observed effects. Mental health, smoking, and alcohol abuse also did not explain our findings. Risk factors for olfactory loss (male gender, lower socioeconomic status, BMI) were included in our analyses, and though they replicated prior work [41], did not affect our results.” Note that the study did not control for effects of possible episodic exposure to toxins or injury that may result in temporary or permanent olfactory impairment not related to death.

Olfactory response is an involuntary response to a stimulus, and may be considered a reflex action. In the human study, presence or absence of smell detection for rose, leather, orange, fish, and peppermint were summed and related to delayed mortality. Olfactory responses to various substances can be scored as present or absent and summed to predict delayed mortality. In the same way, the RAMP method is an example of presence-absence scoring with summation of reflex impairment and injury scores to predict delayed mortality.  Measuring and summing whole animal responses, i.e., olfaction, reflex actions, and injury to stimuli is a powerful method for observing the effects of stressors and aging on delayed mortality.   

“We believe olfaction is the canary in the coal mine of human health, not that its decline directly causes death. Olfactory dysfunction is a harbinger of either fundamental mechanisms of aging, environmental exposure, or interactions between the two. Unique among the senses, the olfactory system depends on stem cell turnover, and thus may serve as an indicator of deterioration in age-related regenerative capacity more broadly or as a marker of physiologic repair function [13].”

Clearly, measurement and summation of presence-absence for whole animal involuntary characteristics (olfaction, reflex actions, and injury) is a powerful way to predict delayed mortality in humans and other animals.

Flexibility of using RAMP to determine bycatch mortality rates for Tanner crab caught in Alaska bottom trawls

Tanner crab (Chionoecetes bairdi), AFSC

Yochum et al. 2015 evaluated the flexibility of RAMP methodology by creating a RAMP for Tanner crab (Chionoecetes bairdi) discarded from the groundfish bottom trawl fishery in the Gulf of Alaska and comparing it to a previously established RAMP for unobserved Tanner crab bycatch (encountered gear and remained on the seafloor) from the bottom trawl fishery in the Bering Sea. The authors found that: “The two RAMPs and the overall mortality rates calculated using these predictors were comparable. However, we detected significant differences between RAMPs. While probabilities of mortality were similar between the two studies for crab with all or no reflexes missing, discarded crab with intermediate reflex impairment had lower mortality probabilities than those from the unobserved-bycatch study. Our results indicate that a RAMP may produce more accurate mortality estimates when applied to animals experiencing similar stressors as those evaluated to create the RAMP, through similar methodology.”

Conditions for holding crab after exposure to stressors can control delayed mortality and should be considered in experiments. “There were differential mortality rates by holding type. Higher mortality rates occurred in the on-board tanks (where the crab were held for the first few days) and in the laboratory tank. Moreover, Score-zero crab died in the holding tanks, but not in the at-sea cages. These results indicate that holding tanks contribute additional stressors, either due to transport, additional handling, or stress from being held in an unnatural setting or at temperatures greater than what was experienced in their natural environment. 

Our holding duration of two weeks was sufficient to determine mortality for all Scores. Given that it can take longer for Score-zero animals to die than those with higher Scores, our holding period allowed us to sufficiently capture Score-zero mortalities. However, the death of a Score-zero crab at day 12 may indicate that holding for more than a week confuses mortality attributed to fishing stressors with that from captivity.”

Evaluation of RAMP flexibility was made by comparing results from different studies. “To evaluate the divergence between the RAMPs we analyzed the differences between the studies. The primary difference was in experimental methods, namely the treatment of the crab before assessment. Crab from the Discard-mortality study were exposed to air for 90min on average (range from 9 to 230min) without any “recovery” in water. In contrast, crab from the Unobserved-mortality study had only brief air exposure and were held in water while awaiting assessment (generally less than 15 min), which may have allowed some recovery. 

These differences in air exposure and recovery in water probably affected the relationship between observed reflex impairments and delayed mortality and hence accounted for the discrepancy between RAMPs. Prolonged air exposure and experiencing cold temperatures was linked with increased delayed and instant mortality, number of autonomies for crab, as well as reduced vigor, juvenile growth, and feeding rates (Carls and O’Clair, 1995; Giomi et al., 2008; Grant, 2003; Stoner, 2009; Warrenchuk and Shirley, 2002). Stoner (2009) found that reflex impairment score and exposure to freezing temperatures were nearly linearly related for Tanner crab. Moreover, he found that the different RAMP reflexes had variable sensitivity to freezing temperatures, namely that the chela closure reflex was the most sensitive reflex, and mouth closure was least. Similarly, Van Tamelen (2005) found that the legs and eyes of snow crab cooled faster than the body, perhaps making them more susceptible to cold air exposure. We hypothesize that the prolonged air exposure for the Discard-mortality study likely impaired the crabs’ reflexes and resulted in higher Scores.”

Recommendations made by the authors. “Results from this study indicate that bias can be introduced in mortality rate estimates when using a RAMP created for one study to estimate mortality rates for a different study where the experimental methods differ, especially with respect to air exposure and recovery in water before assessment. However, when RAMP is used only to approximate mortality rates or to make comparisons between gear types or uses, a previously established RAMP could be used with caution, especially if animals with intermediate Scores are not predominant. For more accurate bycatch mortality rate estimates, our results indicate the importance of using a RAMP that was created by assessing animals that experienced similar stressors to those which the RAMP will be applied. Namely, the procedure for assessing the animals should be similar. We feel that the amount of time the animal spends out of water before assessment be standardized within a time range, along with whether or not the animal is allowed to recover in water before assessment, unless these variables are the treatments being studied.” 

“Our results indicate that consistency in methodology and relevance with respect to mimicking actual fishing stresses for the RAMP approach increases the flexibility of RAMP. It is therefore important, when creating a RAMP, to create repeatable methods that are well documented when publishing. RAMP reflexes should be assessed in a specified order to prevent bias from reflexes that are physiologically linked. If there is a reflex that influences the determination of other reflexes it should be assessed last or not at all. Reflexes that are difficult to determine presence or absence should not be used, and it should be clear in the methods what constitutes an “absent” reflex and how immediate mortalities are treated (are they given a Score or classified separately?). In addition, when a RAMP is being created, data should be recorded on all possible stressors, including injury, and evaluated for their contribution to mortality. Moreover, effort should be made (within the logistical constraints of field and laboratory research) to minimize additional stressors that are unrelated to the fishing stressors of interest.”

Sublethal effects of simulated angling capture (fatigue and air exposure) in snapper: reflex impairment and physiological stress.

Snapper, Pagrus auratus. Floor Anthoni 2006.

A study of reflex impairment and physiological stress was conducted with captured snapper and published in JEMBE 2014 by McArley, T.J & Herbert, N.A.  Fish were exposed to simulated angling by chasing to fatigue, followed by air exposure.  The authors' text is quoted below.

Reflex impairment (RI) was measured for seven reflex actions (Table 1). “If the presence of a positive reflex response was ambiguous it was scored as absent. The entire RI assessment was completed in less than 50 s, of which the fish was exposed to air for approximately 30 s. Reflexes were scored present (1) or absent (0) for individual fish and the RI score (proportion of reflexes impaired) was calculated by dividing the number of reflexes absent by the total number of possible reflexes (Davis, 2010). For example if four out of the seven possible reflexes were absent a fish would be given an RI score of 0.57.”

Reflex impairment is a potential measure of vitality loss after exposure of snapper to angling stressors: 

“A primary aim of this study was to assess the potential use of RI as a simple tool for measurement of fish vitality following angling and our lab-based trials indicate RI has the potential to be used in this way. RI was significantly related to the duration of strenuous exercise and air exposure (Fig. 1) and therefore provided a good index of fish condition. Importantly, fish exposed to more severe stress treatments exhibited greater RI than those exposed to more mild stress treatments, a finding that agrees with several other studies of RI in fish (Barkley and Cadrin, 2012; Brownscombe et al., 2013; Campbell et al., 2010b; Davis, 2007; Humborstad et al., 2009; Raby et al., 2012, 2013). Furthermore, greater RI was associated with significantly higher plasma lactate concentration and reduced muscle pH suggesting that RI can indicate (predict) an alteration in physiological condition.” 

Anaerobic respiration is associated with lactate production and reflex impairment:

“Burst swimming is powered by anaerobic respiration fueled by stored energy in white muscle (Milligan, 1996) and the lactic acid produced accumulates rapidly in muscle tissue and then “spills over” into circulation after a 5–10 min delay (Wood, 1991). Plasma lactate therefore serves as a useful indicator of anaerobic respiration in fish (Gale et al., 2011; Lowe and Wells, 1996; Meka and McCormick, 2005) and, as fish performed burst swimming during chasing, it is unsurprising that plasma lactate correlated positively with the duration of chasing stress and that both muscle pH and blood pH were lower in fish chased for longer periods. Physiological alterations appeared to be more pronounced in summer than in winter suggesting that when water temperature is warmer a more severe stress response appears to occur in snapper.”

Physiological basis for reflex impairment: 

“RI is thought to have a physiological basis (Davis, 2010) and a significant relationship between RI and increased plasma lactate concentration has been observed in salmonids (Raby et al., 2013). As RI is measured directly after stressor exposure physiological disturbances that manifest quickly during stress are likely causes of RI. Physiological alterations such as cortisol concentration that can plateau 30 min - 1 h after the initial stressor exposure (Milligan, 1996; Wendelaar Bonga, 1997) are therefore unlikely to be directly responsible for the RI measured in this study. In this study most RI occurred as a result of an inability to perform reflexes involving powerful muscular contractions, such as the gag reflex, body flexing and the startle response. Powerful muscular contraction is fueled by anaerobic metabolism in white muscle fibres and can only be maintained for short periods (Milligan, 1996). As higher RI scores were correlated with lower muscle pH and higher concentrations of plasma lactate it is hypothesized that muscle fatigue resulting from anaerobic metabolism performed during strenuous exercise caused the majority of the observed RI. The muscle pH and plasma lactate concentrations associated with the same RI scores, however, were different in summer and winter (Fig. 4) and there was no difference between the summer and winter measures of fish vitality (RI) and mortality. This suggests that rather than being causes of RI, plasma lactate concentration and muscle pH may have been indicators of an unmeasured physiological process that impaired some of the reflexes quantified in the current study. Other reflexes we measured that were not as commonly impaired, such as the righting response and vestibular ocular, are essentially neurological and their impairment likely results from alternative pathways to those measured in this study.”

Mortality was rarely observed when snapper were exposed to angling conditions: 

“Despite the limitations of comparing our mortality estimates to real fishing scenarios the findings provide evidence that strenuous exercise and air exposure imposed during angling, are not likely to be direct causes of discard mortality in P. auratus. During the collection of sub-legal snapper from the wild for this study, fish were landed relatively quickly (approximately 15 to 30 s), and were typically unhooked in less than 30 s. In investigations of authentic angling events for P. auratus of comparable size (< 270 mm FL) in south eastern Australia, the majority of fish were landed in less than 30 s (Broadhurst et al., 2012; Grixti et al., 2010) and had less than 30 second exposure to air (Broadhurst et al., 2012). Thus, the 5 minute strenuous exercise period and the 3 minute air exposure period in this study must be considered extreme levels for recreational angling and probably rarely occur in authentic angling events. Encouragingly, even with these high levels of stress, little mortality was seen against a backdrop of high summer water temperatures.”

Predation risk for snapper that show reflex impairment associated with angling: 

“It is often overlooked in catch and release studies but a potentially important contributor to discard mortality is post-release predation (Raby et al., 2014). While no measure of predation risk was assessed in this study, our measurements of RI indicate that snapper may not be overtly susceptible to predation upon release, at least when no barotrauma is present. This is because RI was minimal among snapper released after angling simulations most relevant to authentic recreational angling events (i.e. 0.5 min chasing with up to 1 min air exposure), and it is believed that the vigorous condition of these fish would not make them easy targets for predators. Importantly, reflexes that might be associated with reduced predator avoidance, such as the startle response and righting response, remained intact.”

Snapper captured by trawl may be at risk from increased air exposure: 

“It is likely that mortality increases significantly at some point beyond 3 min air exposure in P. auratus but this may not be relevant to recreational angling. Longer periods of air exposure, however, may be present in commercial trawl fisheries where large catches are sorted on deck so knowledge of air exposure tolerance beyond that observed in this study would be useful in this context. Therefore, the existence of a predictive relationship between RI and mortality in snapper remains a possibility but probably requires the inclusion of more extreme air exposure treatment to be clarified in future trials.”

Clearly any capture of snapper that produces barotrauma can be a source of mortality and requires further study in deep water commercial and recreational fisheries.

Making and Using RAMP in Fisheries

A video is available that explains making and using RAMP in fisheries.

Why is vitality impairment related to mortality?

By definition, healthy animals have full vitality. Vitality becomes impaired as animals become stressed by capture and handling. Severe vitality impairment can result from the effects of physical injury or other stressors, e.g., fatigue, temperature, light, sea state, and air exposure. Maladaptive stress responses or critical injury associated with severe vitality impairment can result in immediate and delayed mortality.

Why score reflex actions and injury?

Reflex actions are fixed behavior patterns that are directly related to vitality impairment, without control by volitional behavior factors, e.g., motivation, hunger, fear, shelter seeking, migration, and reproduction. Reflex actions reflect the state of neural, muscle, and organ functions.

Injuries are directly related to vitality impairment because they can control neural, muscle, and organ functions.

Scoring vitality impairment in general

Any type of reflex action or injury that is related to vitality can be summed to score vitality impairment. The important point is that a sum of presence/ absence scores for vitality characteristics produces an index of vitality impairment. This vitality index can then be used as a measure of variability for sublethal stressor effects in fisheries, as well as a validated indicator and predictor of mortality and survival.

Steps for making and using RAMP in fisheries.

Ecological significance of cold shock: reflex action impairment in bonefish

Bonefish, Florida Sportsman

Fast moving weather fronts or upwelling events can rapidly drop water temperature in sub-tropical areas. Effects of cold shock were studied in bonefish by Szekeres et al. 2014. Fish at 25^oC were exposed to either 18^oC or 11^oC for 2 hours. Ventilation rate and reflex actions were monitored throughout the cold shock. Five reflex actions were tested before and after cold shock, including equilibrium, body flex, vestibular-ocular response, tail grab, and head complex (Brownscombe et al. 2013). Given that the focus of this study was on sub-lethal effects, cold shock exposure was terminated if the fish lost equilibrium. Blood plasma and swimming ability, defined as line crossings and time to loss of equilibrium associated with chasing were also sampled during the experiments.

The authors found that “Behavioral responses of bonefish to cold shock were generally characterized by decreased ventilation rates for the 7°C below ambient treatment with little reflex impairment, and extreme behavioral and reflex impairment in the 14°C below ambient treatment. Fish in the latter treatment exhibited varying periods of hyperactivity followed by impaired or no swimming ability, reduced responsiveness, and the loss of equilibrium, which are all common traits of cold shock exposures.” Experiments with bonefish exposed to the 14°C below ambient temperature were terminated after 30 minutes, as fish lost equilibrium.

Importantly, the authors found “Despite the fact that bonefish in the 14°C below ambient treatment had almost complete reflex impairment during the exposure and sustained high blood lactate concentrations than other treatments, post-exposure swimming abilities were similar to handled control fish. This suggests that although fish become highly behaviorally impaired at colder temperatures, if they are able to escape to more suitable conditions, swimming abilities quickly return and they are unlikely to experience further fitness consequences due to behavioral impairment (e.g. higher predation risk).” 

There “are many facets that have yet to be explored as this research was the first attempt to understand the sub-lethal consequences of cold shock on these sub-tropical fish species. Our research only considered swimming ability as a proxy to understand predation risk in the wild. Future research may focus on determining whether the fish experience compromised disease resistance, poor foraging decisions, changes to fecundity or altered developmental stages. The combination of a changing climate and the economic importance of bonefish throughout the Caribbean warrant more research to be conducted on this species and their responses to an array of changes to ambient conditions.”

Methods for estimating discard survival in fisheries: an integrated approach

Discarding Pacific halibut, FAO

ICES has published a report on methods for estimating discard survival in fisheries. The report details the results of the February, 2014 ICES WKMEDS workshop on discard survival.

“This report will:

-  describe the concepts behind assessing discard survival (Sections 2 and 3);

-  describe three different approaches for estimating survival (vitality assessment, captive observation and tagging) (Sections 4, 5 and 6); and 

-  provide guidance on the selection of the most appropriate approaches and experimental designs, as well as how to integrate and utilize information from them, with respect to specific discard survival objectives (Sections 3, 7, 8 and 9). 

Later versions of this report will cover in more detail: 

-  techniques for assessing survival using tagging and biotelemetry; and 

-  the most appropriate methods for analyzing and reporting survival data. 

It is assumed that the user of these guidance notes has sufficient scientific training, or at least access to suitable scientific support, to be able to conduct the techniques described in these notes in an appropriately systematic and disciplined manner. However, these guidance notes are intended also to be informative for other stakeholders associated with fishing (primarily fishers and managers) who wish to support and understand discard survival estimates.”

The ICES WKMEDS report is a summary of an integrated approach for estimating discard survival. The approach uses various combinations of vitality assessment, captive observation, and tagging to achieve realistic estimates for discard survival in fisheries. The combinations of methods are determined by scientists, stakeholders, and managers using evaluation and prioritization:

“the choice of which species in which fisheries to study depends upon several criteria: existing survival information, the biological traits of the species, its population status, magnitude of discarding, fishery characteristics, environmental characteristics, socio-economic value of the fishery, available resources, and management policy. The process of prioritizing is unlikely to be simple and may involve a number of iterations, where results of preliminary studies inform the final choice.”

The ICES WKMEDS report represents a new approach for estimating discard survival. Sources of information about objectives, priorities, resource implications, and time frames are included in a decision matrix. Managers can use the matrix to make informed choices about discarding in key fisheries and management units and what methods can be used for further study of discard survival. Initial calibration of vitality assessment using delayed mortality observations of discards creates validated indicators for survival. Then use of validated vitality assessment indicators such as RAMP (Reflex Action Mortality Predictors) can provide rapid real-time assessment of potential discard mortality on-board fishing vessels.

ICES. 2014. Report of the Workshop on Methods for Estimating Discard Survival (WKMEDS), 17–21 February 2014, ICES HQ, Copenhagen, Denmark. ICES CM 2014/ACOM:51. 114 pp.

Cautionary tale of rockfish barotrauma and survival: looks can be deceiving

 Yelloweye rockfish, ADFG

Canary rockfish, WDFW

Canary and yelloweye rockfish were captured by Hannah et al. 2014 at 46-174 m depth, retrieved to the surface, and then submerged to depth in specialized sea cages for evaluation of survival.

The authors state, “The external physical signs associated with extreme expansion and retention of swimbladder gas (pronounced barotrauma), including esophageal eversion, exophthalmia and ocular emphysema, were common for both species at these capture depths and were more frequent than in prior studies conducted at shallower depths. Despite similar frequencies of most external barotrauma signs, 48-h post-recompression survival of the two species diverged markedly as capture depth increased. Survival of yelloweye rockfish was above 80% across all capture depths, while survival of canary rockfish was lower, declining sharply to just 25% at capture depths greater than 135 m. Fish of both species that were alive after 48 h of caging displayed very few of the external signs of pronounced barotrauma and had a high submergence success rate when released at the surface.”

Survival and submergence success of canary and yelloweye rockfish, Hannah et al. 2014

Difficulty for evaluating vitality and potential survival by observing barotrauma symptoms and reflex actions is outlined by the authors. “The divergence of 48-h post-recompression survival of canary and yelloweye rockfish as depth of capture increased beyond 135 m shows how difficult it can be to evaluate the survival potential of rockfish with barotrauma based on their appearance at the surface. Most specimens of both species captured at these depths showed some signs of pronounced barotrauma, yet nearly all of the yelloweye rockfish survived following recompression while many of the canary rockfish perished as capture depth increased beyond about 75 m. Studies of post-recompression release behavior also support the notion that surface observations are not indicative of survival, at least for rockfish that tend to retain most of their expanded swimbladder gas (Hannah and Matteson, 2007; Hannah et al., 2008a). The retained gas can make it very difficult or impossible for rockfish to submerge (Hannah et al., 2008b; Hochhalter, 2012) and interferes with the evaluation of reflex behaviors, which have been shown to be useful predictors of survival in other captured and discarded fishes (Davis, 2007; Davis and Ottmar, 2006).”

With regards to stock management, the authors state, “The estimates developed in this study can be very useful for informing the management of hook-and-line fisheries that encounter these two overfished species, especially in combination with data on submergence success as a function of capture depth, like that provided by Hochhalter (2012) for yelloweye rockfish. For example, a primary recommendation from prior studies of post-recompression survival and submergence success for these two species was that hook-and-line fishers should use a variety of “descending” devices to help released fish overcome surface buoyancy (Theberge and Parker, 2005; Hochhalter and Reed, 2011; Hannah et al., 2012; Hochhalter, 2012). The data from this study suggest that descending devices may have a positive effect on survival of yelloweye rockfish across a wide depth range (Fig. 6, lower panel). However, for canary rockfish captured at depths greater than 135 m, survival may be so low that it might be better to either allow retention of these fish or to simply not allow a fishery to operate at these deeper depths (Fig. 6, upper panel).”

The importance of vitality in fishing experiments

Key fishing stressor factors, Davis, 2002

Knowledge of key factors controlling fisheries is necessary for sustainable management of fishery stocks. Scientific hypothesis testing in the form of fishing experiments is a necessary component of fisheries knowledge development and validation. Fishing experiments are performed in the field by simulating actual fishing conditions, by actual fishing, and during survey cruises. Fishing experiments can be used to identify key stressor factors that control and contribute to the survival and mortality of captured, discarded, or escaped animals as well as identifying the key factors controlling fishing gear capture efficiency and selectivity.

Trawl captured animals, Robert A. Pawlowski, NOAA Corps

While field fishing experiments represent realistic conditions, they are a matrix of confounded factors which cannot be easily separated into mechanistic hypothesis tests and explanations of factor importance. Effects of factors are often synergistic and prior animal stressor history can alter relative effects of subsequent exposure to factors, e.g., depth changes, injury, elevated temperature, air exposure, and size and species differences.

Flow chart of experimental fishing stressor factors, Davis and Olla 2001

Simulated fishing experiments with factors in controlled laboratory conditions is one way to test hypotheses about mechanistic effects of individual factors and their interactions. However these laboratory experiments are generally viewed as not realistic to field conditions and they are used to identify factors that may be important in the field. Furthermore, modern requirements of animal care laws and committees restrict the use of laboratory fishing experiments by not allowing human application of experimental stressor factors on animals and the use of mortality outcomes. These same laws and committees do not have jurisdiction over field fishing experiments. 

Laboratory trawl tow tank, NOAA RACE

Given that factors are confounded in field fishing experiments, how can we test for effects of factors in the traditional mechanistic hypothesis test? We can test for changes in animal vitality. Since vitality has been shown to be correlated with survival and mortality, it is a useful indicator of animal outcomes before and after exposure to experimental stressor factors. For example, we generally do not know the exposure of animals to stressors prior to experimental manipulation of factors. Not knowing the complete stressor profile is not an obstacle since the animal knows the complete stressor profile and presents vitality levels that have integrated the effects of that profile. Then we can expose animals to additional stressor factors and measure further changes in vitality from their initial levels. 

Important to shift mechanistic thinking from needing to know the effects of individual factors to knowing the effects of fishing variability. Manipulations of time in air and elevated temperature represent differences in fisher sorting and handling behavior on deck and are appropriate for defining the effects of fishing variability. Effects of variation in tow time and catch quantity can be manipulated and are included in the mix of animals landed. The questions of associations among individual fishing stressor factors is left for another day and are more of interest to mechanistic scientists than to managers and fishers. Fishing variability will give a picture of the fishery and its potential effects on animal vitality. By measuring animal vitality, which integrates the effects of stressor factors, you have measured a key master variable that indicates the important effects of fishing.

Vitality is the key variable that can be used to indicate and predict delayed survival and mortality outcomes for discards and escapees from fishing. The relationships between vitality and survival and mortality are defined by captive observation or tagging and biotelemetry experiments. During exposure of animals it is important to insure that all stressor types normally in the fishery in question are present for the population of tested animals (e.g., temperature, air exposure, fatigue, injury) and that a full range (0-100%) of vitality impairment and mortality are observed. Then relationships can be calculated for each species of interest that do not extrapolate beyond available data ranges and that apply to the fishery of interest. These relationships can then be used to predict survival and mortality for animals under any condition of interest in the fishery without the need for further captive observations or tagging.

Consider how scientific peer-reviewers may see this shift from mechanistic thinking and develop thoughts that elaborate the importance of vitality from the animal’s point of view. Some resistance is expected from mechanists who believe that they can attribute cause and effect to individual factors. There is always a matrix of interactions, even under the most restrictive and controlled experimental conditions. There are always interactions and synergisms to account for. As a result, there are associations among factors, rather than cause and effect. In other words, there are causes and conditions associated with effects. 

From the fishing experiment perspective, we set up fishing conditions that are real or that simulate fishing and then measure animal vitality, which is an integrated measure of the effects of interacting factors. It is useful to identify the important stressors by experimentally changing them in fishing experiments; changes in time in air, trawl time, trap retrieval time, depth, season, temperature, catch amount, and injuries. Always remember that there is a hidden context of conditions, i.e., the animals are prestressed by other factors not being controlled. But this hidden context can be accounted for by observing and comparing vitality impairment among animals observed in all treatments, including simply captured animals without additional stressor exposures (using positive controls). This experimental approach is useful both for fishers who wish to modify fishing gear and practices, as well as managers who wish to observe animal vitality and correlate that with mortality and survival.