Indicator Of Stress In Fish Research Articles

Assessment of fish health status for waterless transportation based on image features and deep learning models

In the current aquaculture industry, the detection of physiological stress indicators in fish during waterless transportation is considered to be an effective method to monitor their health status. However, commonly used detection methods are invasive and interfere with the accuracy of detection results. In order to solve this problem, this study takes grouper as an example, combines multi-sensor physiological stress indicators monitoring, and designs a health status assessment and classification model based on image features and deep learning models, which inputs real-time fish image feature information data acquired during low-temperature waterless transportation into the optimised CNN-BiGRU classification model based on whale optimisation algorithm (WOA) proposed in this study, so as to achieve an accurate assessment of the health status of the fish. By inputting the image feature datasets based on time series and classified into the WOA-CNN-BiGRU, BiGRU, and GRU classification models for training, validation, and testing, respectively, the effectiveness of the classification performance of the models was verified by using accuracy, precision, recall and F1-value composite scores as the metrics. It was found that the WOA-CNN-BiGRU model had excellent classification performance, followed by the BiGRU and GRU models, especially in classifying the health status assessment of small size grouper. This study proposes an efficient and inexpensive method for real-time monitoring and accurate assessment of the health status of fish during waterless transportation, and also provides a reference for vigour monitoring and health assessment of other similar aquatic products.

Effects of chlorpyriphos on haematological parameters in freshwater fish Anabas testudineus

Haematological measures are frequently employed as stress indicators in fish and to evaluate their overall health. This study evaluated the sublethal effects of the organophosphate insecticide chlorpyriphos on a number of haematological parameters in freshwater fish, specifically Anabas testudineus. Based on comparisons between control and experimental groups exposed to sublethal amounts of chlorpyrifos (0.125 and 0.250) at 7, 14, and 21 days, the effect was evaluated. Red blood cell counts, haemoglobin levels, WBC, and thrombocyte counts were found to decline as chlorpyrifos concentrations and days increased. White blood cell counts rose on the seventh day of exposure but fell on the fourteenth and twenty-first days when chlorpyrifos concentrations rose. According to the study, chlorpyrifos had a major negative effect on the fish's health, and haematological measures might be used as a diagnostic tool to determine whether A. testudineus has been exposed to chlorpyrifos.. KEYWORDS :A. testudineus, Chlorpyrifos, haemoglobin, Rbc, Wbc, haematological.

PARASITIC FAUNA OF MALAPTERURUS ELECTRICUS AND PARACHANNA OBSCURA FROM LEKKI LAGOON, LAGOS, NIGERIA

Host condition factor and gastrointestinal parasites in Malapterurus electricus and Parachanna obscura in Lekki lagoon, Lagos, Nigeria were investigated from September, 2014 to February, 2015. Eighty (80) fishes of Malapterurus electricus and eighty (80) of Parachanna obscura were caught and dissected for intestinal helminth parasites and protozoa. Condition factors of all individuals were determined. Median condition (K < 2.17 and K > 2.17) for Malapterurus electricus, (K < 1.02 and K > 1.02) for Parachanna obscura were used in grouping the individuals. Protozoa infection in the gut includes; myxosporidia and coccidians. Heterogeneous infection of these protozoa in low condition Parachanna obscura were; Myxosporidia, 0.20, Coccidia, 0.10 and high condition individuals, 0.10, 0.15 respectively. There were no coccidian infection in M. electricus, but myxosporidia in low and high condition individuals were 0.15 and 0.10. The rate of infection of Electrotaenia malopteruri, a cestode, specific to M. electricus was higher among high condition individuals, 0.15, compared low condition individuals, 0.10. Procamallanus sp, had the highest prevalence among the high condition individuals of Parachanna obscura, the trend was; High Condition M.electricus (0.10) < Low Condition P.obscura (0.15) < Low Condition M.electricus (0.20) < High Condition P.obscura (0.25). Infected individuals of low condition factor (K < 2.17) of Malapterurus electricus had a greater histopathological alteration index (HAI, 18.0) compared to the high condition individuals (HAI, 8.0). These individuals had from mild to severe ulceration of the mucosa and congestion of the blood vessels. In infected individuals of low condition factor (K < 1.02) of Parachanna obscura, there were mucosal edema and haemorrhage in their intestinal walls. They also had greater histopathological alteration index (HAI, 22.0) compared to the high condition individuals (HAI, 6.0). Prevalence of gastrointestinal parasitic fauna in low condition host could be an indicator of stress in fish

Measuring cortisol in fish scales to study stress in wild tropical tuna

Cortisol is recognized as a physiological indicator of stress in fish. However, this hormone is typically measured in plasma samples. In this study, cortisol content was evaluated for the first time in the scales of tropical tuna. The sample included 20 skipjack tuna (Katsuwonus pelamis) and 25 yellowfin tuna (Thunnus albacares) collected in the Atlantic (the Gulf of Guinea off the Ivory Coast) and the Indian Ocean (off Reunion Island), and their scales were analyzed using an ELISA test. The results indicated that on average, cortisol concentration (1) did not show any geographical pattern, (2) was independent of fish size, and (3) was significantly higher in skipjack (mean 4.75 ± 6.56 ng g−1) than in yellowfin (mean 1.65 ± 1.85 ng g−1), although the difference was mainly due to four skipjack individuals. Larger datasets would be needed to confirm any species difference in cortisol concentration. The particularly high cortisol concentration observed in four individuals may be due to fish from different schools, suggesting the need for future sampling from free-swimming schools to investigate intra- and inter-school variability in cortisol concentration. This study opens the door for future research, including the collection of scales on tagged individuals, to investigate the links between chronic stress and behavior in these species.

Comparative assessment of cortisol in plasma, skin mucus and scales as a measure of the hypothalamic-pituitary-interrenal axis activity in fish

Cortisol, the end product of the hypothalamus-pituitary-interrenal (HPI) axis, has been traditionally measured in blood as indicator of stress in fish, however, the degree of invasiveness inherent to blood collection is not always possible or desirable. Instead, cortisol measurement in skin mucus is far less invasive, but as blood, this method potentially provides only a brief window of information of the HPI axis activity. The newly described method of cortisol measurement from scales may serve as a long-term, integrated measure of the HPI axis activity in fish. While skin mucus and scales cortisol measurement present practical and conceptual advantages, there are still several unclear issues related to their biological relevance that need deeper study. Accordingly, we aimed to evaluate whether skin mucus and scales cortisol levels can be reliably used as stress indicators by subjecting fish to prolonged, continuous stressful conditions. The present study demonstrates that the measurement of cortisol in skin mucus reflect circulating cortisol concentrations when fish are responding to stress with an intense activation of the HPI axis. Results also revealed that cortisol content in scales strongly correlates to circulating cortisol levels in chronically stressed fish. Besides, we provide further support that scales cortisol assessment offer a retrospective measure of the past stress experience in fish. While this study provides a good basis for future research applying the methods presented, our results open the question of whether these matrices have additional sources of cortisol other than blood, and the route of incorporation or diffusion. Further knowledge about the general robustness and stability of scales cortisol in fish subjected to prolonged stress would largely help strengthen the interpretation of hormone fluctuations in this matrix.

Changes in natural haemolytic complement activity induced by stress in gilthead seabream (Sparus aurata L.)

In aquaculture, animals can be continually exposed to environmental stress factors that put their health and even survival at risk. Two experiments were performed to evaluate the impact of different stress conditions (acute crowding and anaesthetic) on the natural haemolytic complement activity in serum and skin mucus of gilthead seabream (Sparus aurata L.). In the first experiment, fish were subjected to 10 kg m−3 (low density, control group) and 50 kg m−3 (high density, crowding group) during 2, 24 and 48 h. In the second experiment, fish were unexposed (control) or exposed to 40 ppm of MS-222 or 5 ppm or 10 ppm of clove oil for 1 h. In fish maintained in acute crowding conditions only an increase of the haemolytic complement activity was observed in the skin mucus after 24 h of exposure. However, a similar statistically significant increase was observed in serum and skin mucus of fish exposed for 1 h to the lowest concentration of clove oil (5 ppm) tested. The results point to a new and alternative way to assess stress in farmed fish by using skin mucus instead of blood serum and confirm that the measurement of natural haemolytic complement activity serves as an indicator of stress in fish.

Development of an optical communication type biosensor for real-time monitoring of fish stress

Expanding our understanding of the physiologic conditions, including stress, that affects fish health will facilitate the development of practices to improve fish health. Toward this aim, we previously developed a radio transmission-type biosensor for real-time monitoring of glucose levels, which are an important indicator of stress in fish. The use of this system is restricted in seawater, however, because seawater greatly attenuates radio waves. Therefore, we designed another transmission method for use in seawater. In the present study, we developed a novel wireless biosensor system using optical technology through the application of a light-emitting diode (LED) as the transmission device, which was attached to the body of the fish (Oreochromis niloticus) and stress was monitored in fish under free-swimming condition. The LED transmitter was used to deliver a digital signal that was based on the output current of the sensor. The transmission efficiency is influenced by the relative position of the optical axis of the LED and receiver. The communication success rate remained stable for approximately 94% of the total monitoring time when the LED was attached to the head of the fish and the receiver was placed above the water tank. The stress response of the fish was also measured by loading a stressor, such as chemical (ammonium nitrogen compounds, pH), physical (strobe flash) and biologic (threat behavior) stressor. Successful real-time monitoring of fish stress indicated that the new biosensor system worked well and is a promising tool for monitoring on fish condition.

Eye darkening as a reliable, easy and inexpensive indicator of stress in fish

We expand the use of eye darkening (ED) to indicate non-social stress in the fish Nile tilapia Oreochromis niloticus (L.). ED is easily estimated, not requiring any sophisticated equipment, and is non-invasive, facilitating the collection of several measures of stress over time. In the current study, we showed the following: (i) high- and low-ED occur spontaneously, indicating different fish reactions to adjustments to a novel environment; (ii) fish confinement or air exposure clearly increases ED (air exposure is a stronger stressor than confinement), and the time to restore basal values indicates the severity of the impact of the stressor on the fish (this response is not affected by period of the day, e.g., morning or afternoon); and (iii) in adults, females were more responsive (slower recovery) to 2-min air exposure than to 30-min confinement.

Food consumption and digestive enzyme activity ofClarias batrachusexposed to various temperatures

The effect of temperature on the food consumption rate and the digestive enzyme activities of Clarias batrachus (80.60 ± 5.34 g) were evaluated. Fish were exposed to six different temperatures of 10, 15, 20, 25, 30 and 35 °C following an acclimation temperature of 25 °C. The rate of temperature change was 2 °C day−1. Highest food consumption was recorded at 25 °C. It gradually reduced with decreasing water temperature. Food consumption rate was significantly (P < 0.05) lower at 10 °C compared with other treatments. Hence, 46.67, 8.20–23.58 and 1.02–6.15% reduced food consumptions were recorded in groups exposed at 10, 15 and 20 °C temperatures, respectively, compared with the 25 °C. The consumption rate was not affected in fish exposed at 30 and 35 °C. Total protease, trypsin and chymotrypsin activities were significantly (P < 0.05) higher in fish exposed at 25 °C compared with others. Lipase activity was significantly (P < 0.05) higher in fish exposed at 30 °C compared with others. Lowest enzyme activities were recorded at 10 °C. Water temperature below 25 °C affected the food consumption and digestive enzyme activities in fish that served as indicators of stress in fish.

Endocrine (plasma cortisol and glucose) and behavioral (locomotor and self-feeding activity) circadian rhythms in Senegalese sole (Solea senegalensis Kaup 1858) exposed to light/dark cycles or constant light

The existence of daily rhythms under light/dark (LD) cycles in plasma cortisol, blood glucose and locomotor and self-feeding activities, as well as their persistence (circadian nature) under constant light (LL), was investigated in Senegalese sole (Solea senegalensis). For the cortisol and glucose rhythms study, 48 soles were equally distributed in 8 tanks and exposed to a 12:12 LD cycle and natural water temperature (experiment 1). After an acclimation period, blood was sampled every 3h until a 24-h cycle was completed. Blood glucose levels were measured immediately after sampling, while plasma cortisol was measured later by ELISA. In experiment 2, the fish were exposed to LL for 11days, and after this period, the same sampling procedure was repeated. For the study of locomotor and self-feeding rhythms (experiment 3), two groups of sole were used: one exposed to LD and the other to LL. Each group was distributed within 3 tanks equipped with infrared photocells for the record of locomotor activity, and self-feeders for feeding behavior characterization. The results revealed a marked oscillation in cortisol concentrations during the daily cycle under LD, with a peak (35.65±3.14ng/ml) in the afternoon (15:00h) and very low levels during the night (5.30±1.09ng/ml). This cortisol rhythm persisted under LL conditions, with lower values (mean cortisol concentration=7.12±1.11ng/ml) and with the peak shifted by 3h. Both rhythms were confirmed by COSINOR analysis (p<0.05). The synchronizing role of temperature and feeding schedule, in addition to light, is also discussed. Diel rhythms of glucose were not evident in LD or LL. As to locomotor and self-feeding activity, a very marked rhythm was observed under LD, with higher activity observed during the night, with acrophases located at 2:14 and 3:37h, respectively. The statistical significance of daily rhythms was confirmed by COSINOR analysis. Under LL, both feeding and locomotor rhythms persisted, with an endogenous period (τ) around 22.5h. In short, our findings described for the first time the existence of circadian cortisol and behavioral circadian rhythms in flat fish. Such results revealed the importance of taking into account the time of day when assessing stress responses and evaluating physiological indicators of stress in fish.

Alternative matrices for cortisol measurement in fish

Plasma cortisol is the most commonly used indicator of stress in fish but, as the blood sampling procedure itself can be a source of stress, it would be helpful to measure cortisol using less invasive matrices. It is also necessary to find alternative matrices as stress indicators in dead fish in which blood sampling is impossible. In the present study, we investigated transport stress in three aquaculture species, European sea bass (Dicentrarchus labrax L.), common carp (Cyprinus carpio L.) and rainbow trout (Oncorhynchus mykiss Walbaum), by cortisol determination (radioimmunoassay) in plasma and other matrices (skin mucus, gut content, lateral muscle and caudal fin). Cortisol significantly increased after transport in all species and matrices, except in the sea bass gut content, where it remained unchanged. The three species responded to transport stress by producing different cortisol levels. In conclusion, the significant correlation found between plasma cortisol and most of the other matrices opens up the possibility of using them to evaluate stress in fish: mucus sampling is a less invasive method than blood sampling, and in addition to muscle and fin sampling, it can be used in postmortem fish.

Species‐specific welfare aspects of the main systems of stunning and killing of farmed Carp

Species‐specific welfare aspects of the main systems of stunning and killing of farmed Carp

Some changes in the haematological parameters of rainbow trout (Oncorhynchus mykiss) exposed to three artificial photoperiod regimes

Haematological parameters are often used as health status and stress indicators in fish. However, information on the effects of artificial photoperiods on these parameters is scarce and ambiguous. The consequences of three different artificial photoperiod regimes [light/day 12 h:12 h (LD 12:12) for 150 days as controls; LD 14:10 and LD 24:0, both for an initial 60 days, and then LD 12:12 for the remaining 90 days] were evaluated in trout. In all fish the haemogram and the erythrocyte production index (EPI) were assessed at days 7, 14, 30 and 60 (phase 1) and at days 90, 120 and 150 (phase 2). At the end of phase 1, photoperiods LD 14:10 and 24:0 induced higher production of immature (late basophilic) erythrocytes (0.06–0.08 × 1012 cells/l, P < 0.05) and elevated EPI (young erythrocytes = 1.6 in LD 14:10, 2.25 in LD 24:0; late basophilic erythrocytes = 4.1–4.9 in LD 14:10, 3.3 in LD 24:0) than in controls (late basophilic erythrocytes = 0.02–0.04 × 1012 cells/l; EPI young erythrocytes = 0.2–1.0; EPI late basophilic erythrocytes = 1.4–2.7). During phase 2, only reduced numbers of lymphocytes and thrombocytes were observed at day 150 in the LD 14:00 group. An increase in the EPI (1.9) of control young erythrocytes was found at day 150.

Effects of Tricaine Methanesulfonate, Hypno, Metomidate, Quinaldine, and Salt on Plasma Cortisol Levels following Acute Stress in Threespot Gourami Trichogaster trichopterus

AbstractBlood plasma cortisol concentration is an indicator of stress in fish, and anesthetics may serve to ameliorate stress and reduce the cortisol response. Previous studies have concentrated on cold‐ and warmwater species; little information exists for tropical fishes. Mortalities are increased after handling stress of threespot gourami Trichogaster trichopterus (Osphronemidae), a commonly cultured tropical ornamental fish. Plasma cortisol levels were evaluated in threespot gourami after a handling stressor and treatment with one of four anesthetics—tricaine methanesulfonate (TMS; 60 mg/L), metomidate (0.8 mg/L), quinaldine (5 mg/L), and Hypno (0.14 mg/L)—or salt (NaCl; 3 g/L). Fish in all anesthetic treatments had significantly lower cortisol levels than either salt‐treated fish or the untreated controls. Therefore, the use of such anesthetics should be beneficial in reducing handling stress. Based on our data, further studies refining the use of anesthetics, particularly metomidate and quinaldine, with tropical ornamental species are warranted. Although salt did not reduce cortisol, it is beneficial as an osmoregulatory aid; therefore, future studies should investigate combination treatments of an anesthetic with salt.

Stress response of red sea bream Pagrus major to acute handling and chronic photoperiod manipulation

A number of primary and secondary features used as stress indicators in fish were investigated in red sea bream, Pagrus major exposed to different photoperiod regimes. In Experiment 1, fish weighing from 17 to 25 g were subjected to acute stress consisted of netting the fish from the rearing tanks and confinement in a small tank (30 L) half-full of water. The fish were then sampled at 0.5, 1, 4, 24, 48, and 72 h post-stress to investigate the stress-induced levels of different parameters. In Experiment 2, fish were exposed to different photoperiods (6L : 6D, 12L : 12D, 16L : 8D and 24L : 0D) for eight weeks. Blood samples were collected from the fish at the 1st, 4th and 8th week after exposure to the photoperiods. Acute stress significantly increased the plasma levels of cortisol (190.1 ng ml − 1 ) and glucose (109.5 mg dl − 1 ) and returned back to the basal levels within 24 h. In contrast, plasma levels of cholesterol (180.5 mg dl − 1 ), total protein (3.0 g dl − 1 ), potassium (4.1 mMol l − 1 ) and Ca ++ (11.3 mg dl − 1 ) were decreased significantly and returned to levels indistinguishable from those of control within 72 h except cholesterol, which did not recover within 72 h. In Experiment 2, fish exposed to 6L : 6D, 16L : 8D and 24L : 0D photoperiods had significantly higher cortisol levels than those exposed to the 12L : 12D photoperiod during the first week ( P < 0.05); however, there was no major difference in cortisol levels among the treatments during other sampling times. There were no significant differences in other parameters among the treatments during the course of the study ( P > 0.05). Fish exposed to the 24L : 0D photoperiod showed significantly higher weight gain and SGR followed by 16L : 8D, 6L : 6D and 12L : 12D photoperiods. The plasma levels of different stress indicators in different photoperiods in Experiment 2 were far lower than those of acute stress-induced levels observed in Experiment 1. On the other hand, no apparent signs of chronic stress were observed in fish exposed to different photoperiods. The results demonstrated that photoperiod manipulation caused no significant acute or chronic stress response in red sea bream.

Caution for using ventilatory frequency as an indicator of stress in fish.

This study tested the use of ventilatory frequency (VF) as an indicator of stress in the Nile tilapia, Oreochromis niloticus (L.). Firstly, we tested the relationship between VF and plasma cortisol after confinement. Confined fish showed higher VF and plasma cortisol levels, but the latter continued to increase significantly for longer time than VF. Secondly, we conducted another experiment to test the use of VF as indicator of fish stress. In four out of six treatment, we confined the fish for different intervals (30 s, 5, 15 or 30 min). The others were used as control. In one, no handling was imposed. The other control consisted of introducing the partition (the same used to perform the confinement) into the aquarium for less than 4 s, without confinement and immediately removing the partition (partition control). Ventilatory frequency was increased for the partition control as much as for the longer duration of confinement. This clearly indicates that VF is a very sensitivity response to disturbance, but of limited use because this parameter does not reflect the severity of the stimulus. Thus, although VF is a non-invasive technique that does not require sophisticated recording equipment, its usefulness is limited.

Sex-related differences in the organismal and cellular stress response in juvenile salmon exposed to treated bleached kraft mill effluent

Exposure of fish to stressors can elicit biochemical and organismal changes at multiple levels of biological organization collectively known as stress responses. The organismal (plasma glucose and cortisol levels) and cellular (hepatic hsp70) stress responses in fish have been studied in several species, but little is known about sex-related differences in these responses. In this study, we exposed sexually immature juvenile chinook salmon (Oncorhynchus tshawytscha) to bleached kraft mill effluent (BKME: 0%, 1%, and 10% v/v) for 30 days and then measured components of their organismal and cellular stress responses. Males exposed to 1% BKME had higher levels of plasma glucose than females. Plasma cortisol levels were unaffected in females exposed to BKME, but males exposed to 10% BKME had significantly higher levels of plasma cortisol relative to non-exposed males. While exposure to BKME did not affect hsp70 levels in males, females exposed to 1% BKME had higher levels of hsp70 relative to non-exposed and 10% BKME groups. Within any given treatment, females had higher levels of hsp70 relative to males. This study demonstrates that sex-related differences exist in commonly used indicators of stress in fish, and points out the importance of considering the sex of the fish in stress research.

Altered stress responses in rainbow trout following a dietary administration of cortisol and β-napthoflavone

Previous studies have demonstrated that fish inhabiting polluted waterways often have an impaired stress response at the organismal level. Given the possible link between the organismal (i.e. cortisol) and cellular (i.e. heat shock proteins; hsp) stress responses, we conducted this study to examine the ability of rainbow trout to respond to a 2 h, +14 °C heat stress (HS) challenge following a 28 d, sub-chronic exposure to increased concentrations of cortisol (5 mg kg−1 b.w.), β-napthoflavone (bnf; 50 mg kg−1 b.w.), and a combination of both (mixture), through the diet (1.5% b.w. every 48 h). While control fish responded to the HS by significantly increasing components of their organismal (cortisol, glucose, and lactate) and cellular (hepatic hsp70 protein) stress responses 6 and 24 h post HS, cortisol-, bnf-, and mixture-fed fish had impaired stress responses at both levels of organization. Additionally, hepatic hsp70 levels were significantly reduced 6 h post HS in cortisol-fed fish. While bnf-fed fish had significantly higher EROD activity, cortisol potentiated EROD activity in the mixture-fed fish. Similarly, plasma cortisol concentrations in the mixture-fed fish were significantly lower relative to cortisol-fed fish. These data are the first to indicate that sub-chronically stressed fish can have impaired stress responses at both the organismal and cellular levels. These findings raise questions regarding: (a) the universal and simple applicability of biological indicators of stress in fish; (b) the possible functional relationship between these two levels of stress responses; and (c) the importance of hsps in the generalized stress response of the whole organism.

Evaluation of Simple Instruments for the Measurement of Blood Glucose and Lactate, and Plasma Protein as Stress Indicators in Fish

Abstract.— Responses of rainbow trout, Oncorhynchus mykiss, to handling and confinement stress were used to validate the efficacy of portable instruments for measuring blood glucose and lactate, and plasma protein. Glucose and lactate increased with time and showed good linear correlation with time after disturbance. The Advantage blood glucose and Accusport™ lactate meters were similarly correlated, except that values were consistently lower than those obtained by established laboratory techniques. Refractometry gave higher plasma protein values than results from biuret reagent but with excellent correlation. These findings indicate a potential role for portable instruments in field and hatchery locations where relative rather than absolute values may be used to evaluate responses to stressors.

Simple field methods for monitoring stress and general condition of fish

While physiological stress in fish can be assessed by several laboratory-based methods, many of those methods use very sensitive and expensive instruments, which make them impracticable under field conditions. The objective of this study was to determine the feasibility of detecting stressed states in fish, under Held conditions, using simple but reliable methods. The first set of experiments were conducted in the laboratory, where a standard handling stressor was applied to juvenile coho salmon, Oncorhynchus kisutch (Walbaum). Results of blood samples analysed by both laboratory-based and portable methods (e.g. glucometers for diabetic patients) were in good agreement. The second set of experiments focused on field-testing the most promising methods based on the laboratatory tests. Production lots of Atlantic salmon. Salmo salar L., at commercial fish farms were sampled to assess stress and health status with the selected field techniques, from the swim-up to smolting stages. The measured variables included: blood glucose, protein, haemoglobin concentrations: haematocrit: erythrocyte and lymphocyte numbers: and an autopsy-based health profile, which involved the internal and external examination of the body for texture and colour. In addition to establishing resting, or normal, values for each variable under field conditions, blood glucose levels were observed to increase significantly at 4 h after a stressful event (e.g. grading, handling) using these field methods. Some of these field techniques were found to be reliable indicators of stress in fish and have potential for use in aquaculture facilities and field monitoring programmes.