The effect of chronic exposure to chloridazon and its degradation product chloridazon-desphenyl on signal crayfish Pacifastacus leniusculus.

The effects of chloridazon (Ch) and its metabolite chloridazon-desphenyl (Ch-D) at the environmentally relevant concentrations of 0.45 µg/L and 2.7 µg/L on signal crayfish Pacifastacus leniusculus were assessed in a 30-day exposure followed by a 15-day depuration period. Locomotion, biochemical haemolymph profile, oxidative and antioxidant parameters, and histopathology were evaluated. Crayfish exposed to Ch at 0.45 µg/L and 2.7 µg/L showed significantly (p < 0.01) higher CAT activity and GSH level in hepatopancreas and gill compared to controls. The concentration of Ch at 2.7 µg/L was associated with significantly (p < 0.01) higher levels of GLU, LACT, ALT, AST in haemolymph compared to controls. Chloridazon-desphenyl exposure at both tested concentrations caused significantly higher (p < 0.01) GLU, LACT, ALT, AST, NH3, and Ca in haemolymph; lipid peroxidation (TBARS) levels in hepatopancreas; and CAT activity and GSH level in hepatopancreas and gill. Alterations of structure including focal dilatation of tubules, increased number of fibrillar cells, and haemocyte infiltration in the interstitium were observed with 2.7 µg/L Ch and with both Ch-D exposures. Locomotion patterns did not vary significantly among groups. A 15-day recovery period was insufficient to restore normal physiological parameters in exposed groups. Chloridazon and its metabolite Ch-D exerts harmful effects on crayfish.

Effects of s-metolachlor on early life stages of marbled crayfish.

The effects of s-metolachlor chronic exposure at concentrations of 1.1 µg/L (maximal real environmental concentration in the Czech Republic), 11 µg/L (environmental relevant concentration) and 110 µg/L on early life stages of marbled crayfish (Procambarus virginalis) was evaluated under laboratory conditions. All s-metolachlor exposures resulted in higher mortality, delay ontogenetic development with accompanied slower growth and excited behaviour (increase of total distance moved and walking speed). Significantly lower superoxide dismutase, catalase, glutathione S-transferase activity and reduced glutathione level was observed at two higher tested concentrations (11 and 110 µg/L) of s-metolachlor compared with the control. S-metolachlor in concentrations 110 µg/L showed alteration of the tubular system of hepatopancreas including focal disintegration of tubular epithelium and notable reduction in epithelial cells number, especially B-cells. In conclusion, potential risk associated with using of s-metolachlor in agriculture, due to effects on non-target aquatic organisms as documented on early life stages of marbled crayfish in this study, should be taken into account.

Effects of chemical cues and prior experience on predator avoidance in crayfish.

Multisensory stimuli provide organisms with information to assess the threat present in the surroundings. Olfactory cues show dominance over other sensory modalities in the aquatic environment. The impact of chemical predator cues combined with experiences gained (learning) in species without previous contact is not fully understood. We investigated the foraging and shelter-seeking behaviour of naïve and experienced marbled crayfish Procambarus virginalis juveniles in response to the chemical signals of pumpkinseed Lepomis gibbosus alone and in combination with alarm chemicals produced by preyed-upon conspecifics. Naïve and experienced (previously exposed to pumpkinseed predation) juveniles were stocked in an arena with shelter and feed and exposed (1) to water from a tank containing a predator actively feeding on conspecifics, (2) water from a tank with predator only and (3) water only as control. Crayfish exposed to the combined stimuli avoided the inlet zone and gravitated to shelter zone of the arena to a greater extent than did those exposed to predator-only cues and the control. Regardless of the treatment, experienced crayfish showed significantly reduced interest in feeding. Our findings imply that crayfish response to threat-associated odours with the greatest potency when visual or tactile cues are present, while previous encounters with predators may make them more cautious.

Effects of praziquantel on common carp embryos and larvae.

This study aimed to assess the toxicity of praziquantel (anthelmintic drug) in different developmental stages of common carp (Cyprinus carpio) based on mortality, early ontogeny, growth, oxidative stress, antioxidant enzymes, histology and behaviour. Praziquantel at all tested concentrations ranging from 1 to 4 mg/L showed no significant adverse effects on mortality, the early ontogeny and behaviour locomotory (activity, moved distance and velocity) of carp after 35-day exposure. Concentrations of 3 and 4 mg/L caused significantly (P < 0.01) lower growth, total superoxide dismutase and catalase activities compared with controls. Praziquantel is safe for the early life of carp in concentrations ≤ 2 mg/L.

Chronic Toxicity of Primary Metabolites of Chloroacetamide and Glyphosate to Early Life Stages of Marbled Crayfish Procambarus virginalis

Degradation products of herbicides, alone and in combination, may affect non-target aquatic organisms via leaching or runoff from the soil. The effects of 50-day exposure of primary metabolites of chloroacetamide herbicide, acetochlor ESA (AE; 4 µg/L), and glyphosate, aminomethylphosphonic acid (AMPA; 4 µg/L), and their combination (AMPA + AE; 4 + 4 µg/L) on mortality, growth, oxidative stress, antioxidant response, behaviour, and gill histology of early life stages of marbled crayfish (Procambarus virginalis) were investigated. While no treatment effects were observed on cumulative mortality or early ontogeny, growth was significantly lower in all exposed groups compared with the control group. Significant superoxide dismutase activity was observed in exposure groups, and significantly higher glutathione S-transferase activity only in the AMPA + AE group. The gill epithelium in AMPA + AE-exposed crayfish showed swelling as well as numerous unidentified fragments in interlamellar space. Velocity and distance moved in crayfish exposed to metabolites did not differ from controls, but increased activity was observed in the AMPA and AE groups. The study reveals the potential risks of glyphosate and acetochlor herbicide usage through their primary metabolites in the early life stages of marbled crayfish.

Comparison of Behavior and Space Use of the European Bullhead Cottus gobio and the Round Goby Neogobius melanostomus in a Simulated Natural Habitat.

The round goby is an invasive fish in Europe and North America that threatens native species by predation and competition. Its habitat preferences are similar to those of the European bullhead, which it displaces from shelters and out-competes for available resources. We assessed the microhabitat preferences, shelter use, and activity of the round goby and European bullhead in single-species experiments in habitat simulator systems to investigate their behavior in a novel environment. Fish were video-recorded for 28 h in the presence of shelter and feed with water velocity ranging from 0.00 to 0.96 m s-1. The two species showed similar behavior under given conditions. A primary difference was in stress-induced behavior in the initial phases of observation. The round goby spent more time in movement when outside the shelter and a longer time in the escape zone in the exploration period during light. Our results confirmed a significant preference of round goby for low velocity areas and a preference for higher velocities in the European bullhead. Both species were able to cope with velocities > 0.7 m s-1. Therefore, the reported invasion success of round goby is probably not driven by space use or activity patterns, but rather by higher adaptability.

A combination of six psychoactive pharmaceuticals at environmental concentrations alter the locomotory behavior of clonal marbled crayfish.

Pharmaceutically active compounds (PhACs) are ubiquitous in the aquatic environment worldwide and considered emerging contaminants. Their effects on growth, behavior, and physiological processes of aquatic organisms have been identified even at very low concentrations. Ecotoxicological investigations have primarily focused on single compound exposure, generally at a range of concentrations. In the natural environment, pollutants seldom occur in isolation, but little is known about the effects and risks of combinations of chemicals. This study aimed to investigate the effects of concurrent exposure to six psychoactive PhACs on locomotory behavior and life history traits of clonal marbled crayfish Procambarus virginalis. Crayfish were exposed to ~1 µg L-1 of the antidepressants sertraline, citalopram, and venlafaxine; the anxiolytic oxazepam; the opioid tramadol; and the widely abused psychostimulant methamphetamine. In the absence of shelter, exposed crayfish moved significantly shorter distances and at lower velocity and showed significantly less activity than controls. With available shelter, exposed crayfish moved significantly more distance, showed higher activity, and spent a significantly more time outside the shelter than controls. Molting, mortality, and spawning frequency did not vary significantly between the groups. Hemolymph glucose level did not vary among groups and was not correlated with observed behaviors. Results suggest that environmental concentrations of the tested compounds in combination can alter the behavior of non-target aquatic organisms as individual exposure of these compounds, which may lead to disruption of ecosystem processes due to their reduced caution in polluted conditions. Further research is needed using varied chemical mixtures, exposure systems, and habitats, considering molecular and physiological processes connected to behavior alterations.

Survival, Growth, and Reproduction: Comparison of Marbled Crayfish with Four Prominent Crayfish Invaders.

Biological invasions are increasingly recognized ecological and economic threats to biodiversity and are projected to increase in the future. Introduced freshwater crayfish in particular are protruding invaders, exerting tremendous impacts on native biodiversity and ecosystem functioning, as exemplified by the North American spiny-cheek, signal and red swamp crayfish as well as the Australian common yabby. The marbled crayfish is among the most outstanding freshwater crayfish invaders due to its parthenogenetic reproduction combined with early maturation and high fecundity. As their introduced ranges expand, their sympatric populations become more frequent. The question of which species and under what circumstances will dominate in their introduced communities is of great interest to biodiversity conservation as it can offer valuable insights for understanding and prioritization of management efforts. In order to examine which of the aforementioned species may be more successful as an invader, we conducted a set of independent trials evaluating survival, growth, claw injury, and reproduction using single-species stocks (intraspecific interactions) and mixed stocks (interspecific interactions) of marbled crayfish vs. other crayfish invaders since the onset of exogenous feeding. In both single and mixed stocks, red swamp crayfish and yabby grew faster than marbled crayfish, while marbled crayfish were superior to both spiny-cheek and signal crayfish in terms of growth. With the exception of signal crayfish, the faster-growing species consistently reached a higher survival rate. The faster-growing species tended to negatively impair smaller counterparts by greater claw injury, delayed maturation, and reduced fecundity. Only marbled crayfish laid eggs as early as 14 weeks in this study, which is earlier than previously reported in the literature. Thus, the success of marbled crayfish among invasive crayfish is significantly driven by relatively fast growth as well as an early and frequent reproduction. These results shed light on how interactions between invasive populations can unfold when their expansion ranges overlap in the wild, thereby contributing to the knowledge base on the complex population dynamics between existing and emerging invasive species.

The effects of the herbicides terbuthylazine and metazachlor at environmental concentration on the burrowing behaviour of red swamp crayfish.

Despite their low concentrations in many aquatic environments, evidence exists to suggest that herbicides do affect non-target organisms. Given that burrowing is a primary life-history trait in crayfish, herbicides could potentially have serious negative effects on these ecologically important freshwater macroinvertebrates. In this study, we exposed the red swamp crayfish Procambarus clarkii to terbuthylazine (a triazine) and metazachlor (a chloroacetanilide) at an environmental concentration of 2.0 µg/L for 28 days, and then observed their burrowing behaviour for two days. The metazachlor-exposed males excavated a greater number of burrows than the other tested groups, with comparable depths and volumes relative to individual specimen weight. The relative depth and volume of female burrows were identical in all groups. The natural habit of female crayfish of constructing deeper burrows than males was marginally significant in the control and META groups but was not significant for relative volume. The hypothesized adverse effects of chronic exposure to real environmental concentrations of herbicides were not documented in terms of either relative depth or volume. However, the increased number of burrows in metazachlor-exposed animals may mean that this invasive species will cause greater damage to embankments and river banks. The mechanisms behind these effects require closer study.

Effects of chloridazon on early life stages of marbled crayfish.

The effects of chloridazon exposure at concentrations of 2.7 µg/L (maximal real environmental concentration in the Czech Republic), 27 µg/L, 135 µg/L and 270 µg/L on early life stages of marbled crayfish (Procambarus virginalis) were evaluated. Significantly higher glutathione S-transferase activity and reduced glutathione level was observed at all tested concentrations of chloridazon compared with the control. Chloridazon in concentrations 27, 135 and 270 µg/L caused delay ontogenetic development and slower growth. Histopathological changes in hepathopancreas were found in two highest tested concentrations (135 µg/L and 270 µg/L). Crayfish behaviour was not altered in control vs. exposed animals, while the activity parameters tend to decline with increasing chloridazon concentrations.

Effects of metazachlor and its major metabolite metazachlor OA on early life stages of marbled crayfish.

The effects of the herbicide metazachlor and its major metabolite metazachlor OA at two concentrations, including environmentally relevant concentrations of metazachlor (0.0115 µmol/l and 0.0790 µmol/l) and metazachlor OA (0.0117 µmol/l and 0.0805 µmol/l), respectively, were evaluated on early ontogeny, growth, behaviour, oxidative stress, antioxidant enzyme levels, histology, and mortality of marbled crayfish Procambarus virginalis. Both tested concentrations of metazachlor and metazachlor OA were associated with significantly lower growth and delayed ontogenetic development compared to controls. Exposure of metazachlor at 0.0115 µmol/l and metazachlor OA at 0.0117 µmol/l and 0.0805 µmol/l resulted in significantly lower activity of total superoxide dismutase (SOD), catalase (CAT), glutathione s-transferase (GST), glutathione reductase (GR), and reduced glutathione (GSH) compared with control and resulted in gill anomalies ranging from wall thinning to focal disintegration of branchial structure. Metazachlor at the environmentally relevant concentration of 0.0790 µmol/l was associated with significant alterations of crayfish distance moved and walking speed. The potential risk associated with metazachlor use in agriculture related to effects on non-target aquatic organisms.

Cardiac and Locomotor Responses to Acute Stress in Signal Crayfish Pacifastacus leniusculus Exposed to Methamphetamine at an Environmentally Relevant Concentration.

Methamphetamine (METH), a central nervous system stimulant used as a recreational drug, is frequently found in surface waters at potentially harmful concentrations. To determine effects of long-term exposure to environmentally relevant levels on nontarget organisms, we analysed cardiac and locomotor responses of signal crayfish Pacifastacus leniusculus to acute stress during a 21-day exposure to METH at 1 µg L-1 followed by 14 days depuration. Heart rate and locomotion were recorded over a period of 30 min before and 30 min after exposure to haemolymph of an injured conspecific four times during METH exposure and four times during the depuration phase. Methamphetamine-exposed crayfish showed a weaker cardiac response to stress than was observed in controls during both exposure and depuration phases. Similarly, methamphetamine-exposed crayfish, during METH exposure, showed lower locomotor reaction poststressor application in contrast to controls. Results indicate biological alterations in crayfish exposed to METH at low concentration level, potentially resulting in a shift in interactions among organisms in natural environment.

Psychoactive compounds at environmental concentration alter burrowing behavior in the freshwater crayfish.

Pharmaceutically active compounds (PhAC) have been increasingly detected in freshwater and marine waterbodies worldwide and are recognized as major emerging micropollutant threat to the aquatic environment. Despite their low concentrations in the environment, there is evidence of effects on non-target aquatic organisms in natural habitats. To assess the potential effects of PhACs on its burrowing behavior, we exposed the red swamp crayfish Procambarus clarkii to methamphetamine or tramadol at the environmentally relevant concentration of 1 µg/L. Methamphetamine-exposed females constructed burrows of lower depth and volume relative to individual weight than did controls. Tramadol-exposed females consistently exhibited a tendency for smaller burrows, but this difference was not significant. Exposed males showed a non-significant tendency to excavate larger burrows compared with the control. Control and tramadol-treated females maintained the natural tendency of constructing relatively deeper and/or larger-volume burrows compared with males. This sex-related pattern was not detected in the methamphetamine group. The rate of human therapeutic PhAC usage is relatively stable year-round, and impacts on crayfish burrowing can be particularly damaging during periods of drought, when the dilution of waste waters is reduced, and burrowing becomes a critical survival strategy. Our results suggest that an increasingly broad range of environmental impacts of PhACs on non-target organisms can be expected in natural ecosystems.

Effects of S-metolachlor and its degradation product metolachlor OA on marbled crayfish (Procambarus virginalis).

Increasing production of energy crops in Europe, mainly maize and rapeseed, has altered patterns of pesticide use in recent decades. The long-term effects of S-metolachlor (S-M) and of its metabolite metolachlor OA (M-OA) at the environmentally relevant concentration of 4.2 µg L-1 and at 42 µg L-1 (ten-fold concentration) on marbled crayfish (Procambarus virginalis) were evaluated in a 28-day exposure and after a subsequent 28-day recovery period. Indicators assessed were behaviour; biochemical haemolymph profile; oxidative and antioxidant parameters of gill, hepatopancreas, and muscle; and histology of hepatopancreas and gill. Results showed biochemical haemolymph profile (lactate, alanine aminotransferase, aspartate aminotransferase, inorganic phosphate), lipid peroxidation in hepatopancreas, and antioxidant parameters (catalase, reduced glutathione, glutathione S-transferase) of hepatopancreas and gill of crayfish exposed to S-M and M-OA to significantly differ from controls (P < 0.01). Antioxidant biomarker levels remained different from controls after a 28-day recovery period. Differences in behaviour including speed of movement and velocity, and histopathological damage to gill and hepatopancreas were associated with S-M and M-OA exposure and persisted after 28 days in S-M- and M-OA-free water. Results suggest harmful effects of low concentrations of S-M and its metabolite M-OA on non-target organisms and provide information for assessing their effects at environmentally relevant concentrations.

Foraging behaviour of top predators mediated by pollution of psychoactive pharmaceuticals and effects on ecosystem stability.

Although pharmaceuticals are recognized as a major threat to aquatic ecosystems worldwide, little is known about their ecological effect on aquatic biota and ecosystems. Drug-induced behaviour changes could have a substantial impact on consumer-resource interactions influencing stability of the community and ecosystem. We combined laboratory experiments and functional response modelling to investigate effects of real wastewater treatment plant (WWTP) effluent, as well as environmentally relevant concentrations of the antidepressants citalopram and opioid pain medication tramadol, on trophic interactions. Our biological system consisted of dragonfly Aeshna cyanea larvae as predator of common carp Cyprinus carpio fry. Exposure to WWTP effluent significantly increased A. cyanea maximum feeding rate, while those parameters in tramadol and citalopram-exposed larvae were significantly lower from unexposed control group. This suggested the potential of all tested pollutants to have an effect on consumer-resource equilibrium in aquatic ecosystems. While WWTP effluent strengthened interaction strength (IS) of consumer-resource interaction dynamics making the food web more vulnerable to fluctuation and destabilization, tramadol and citalopram could inhibit the potential oscillations of the consumer-resource system by weakening the IS. Similar studies to reveal the potential of pervasive pharmaceuticals to change of consumer-resource interactions dynamics are needed, especially when real WWTP effluent consisting of mixture of various pharmaceuticals displayed very different effect from single compounds tested.

Environmentally relevant concentrations of methamphetamine and sertraline modify the behavior and life history traits of an aquatic invertebrate.

Pharmaceutically active compounds are major contaminants of aquatic environments that show direct and indirect effects on aquatic organisms even at low concentrations. The aim of this study was to assess the effects of the illicit drug methamphetamine and the antidepressant sertraline on clonal marbled crayfish Procambarus virginalis. Crayfish exposed to the environmentally relevant concentrations of methamphetamine of ∼1 µg L-1 did not exhibit significant differences from unexposed controls in distance moved, velocity, and activity level with or without available shelter. Sertraline-exposed (∼1 µg L-1) crayfish were significantly more active, regardless of available shelter, and moved greater distances when shelter was available, compared to control crayfish. Crayfish exposed to methamphetamine and sertraline spent significantly more time outside the shelters compared to controls. Sertraline-exposed crayfish spawned more frequently and showed higher mortality than controls. The results suggest that the low environmental concentrations of the tested compounds could alter the behavior and life history traits of crayfish, resulting in higher reproductive effort and mortality.

Chronic toxicity of metolachlor OA on growth, ontogenetic development, antioxidant biomarkers and histopathology of early life stages of marbled crayfish.

The metolachlor OA is a metabolite of herbicide metolachlor and s-metolachlor. The objective of the present study was to assess the effect metolachlor OA on early life stages of marbled crayfish (Procambarus virginalis). The early life stages of marbled crayfish were exposed for 45 days to three concentrations of metolachlor OA: 4.2 µg/L (environmentally relevant concentration, E1), 42 µg/L (E2) and 420 µg/L (E3) under laboratory conditions. The effects were assessed on the basis of mortality, growth, ontogenetic development, behaviour, oxidative stress, antioxidant biomarkers and histopathology. Metolachlor OA caused significantly lower growth, superoxide dismutase, catalase and glutathione s-transferase activity in all tested concentrations. Metolachlor OA in higher concentrations (42 and 420 µg/L) resulted in significantly delayed ontogenetic development, lower reduced glutathione level and lipid peroxidation. Metolachlor OA has not significant effect on behaviour (activity, total distance moved and walking speed). Histological examination revealed alteration of hepatopancreas and gills in crayfish exposed to two higher tested concentrations. Hepatopancreas reflected histomorphological structural changes of individual cell types. Changes of gills included focal hemocytic infiltration together with enlargement of intralamellar space packed with granular substance. In conclusion, chronic metolachlor OA exposure affected growth, ontogenetic development, and the antioxidant system and caused pathological changes in hepatopancreas and gills of early life stages of marbled crayfish.