Metabolic plasticity of mixotrophic algae is key for their persistence in browning environments

Light availability is the main regulator of primary production, shaping photosynthetic communities and their production of ecologically important biomolecules. In freshwater ecosystems, increasing dissolved organic carbon (DOC) concentrations, commonly known as browning, leads to lower light availability and the proliferation of mixotrophic phytoplankton. Here, a mixotrophic algal species (Cryptomonas sp.) was grown under five increasing DOC concentrations to uncover the plastic responses behind the success of mixotrophs in browning environments and their effect in the availability of nutritionally important biomolecules. In addition to the browning treatments, phototrophic, heterotrophic and mixotrophic growth conditions were used as controls. Despite reduced light availability, browning did not impair algal growth compared to phototrophic conditions. Comparative transcriptomics showed that genes related to photosynthesis were down-regulated, whereas phagotrophy gene categories (phagosome, lysosome and endocytosis) were up-regulated along the browning gradient. Stable isotope analysis of phospholipid fractions validated these results, highlighting that the studied mixotroph increases its reliance on heterotrophic processes with browning. Metabolic pathway reconstruction using transcriptomic data suggests that organic carbon is acquired through phagotrophy and used to provide energy in conjunction with photosynthesis. Although metabolic responses to browning were observed, essential fatty acid content was similar between treatments while sterol content was slightly higher upon browning. Together, our results provide a mechanistic model of how a mixotrophic alga responds to browning and how such responses affect the availability of nutritionally essential biomolecules for higher trophic levels.Peer reviewe

Changes in cardiac proteome and metabolome following exposure to the PAHs retene and fluoranthene and their mixture in developing rainbow trout alevins

Exposure to polycyclic aromatic hydrocarbons (PAHs) is known to affect developing organisms. Utilization of different omics-based technologies and approaches could therefore provide a base for the discovery of novel mechanisms of PAH induced development of toxicity. To this aim, we investigated how exposure towards two PAHs with different toxicity mechanisms: retene (an aryl hydrocarbon receptor 2 (Ahr2) agonist), and fluoranthene (a weak Ahr2 agonist and cytochrome P450 inhibitor (Cyp1a)), either alone or as a mixture, affected the cardiac proteome and metabolome in newly hatched rainbow trout alevins (Oncorhynchus mykiss). In total, we identified 65 and 82 differently expressed proteins (DEPs) across all treatments compared to control (DMSO) after 7 and 14 days of exposure. Exposure to fluoranthene altered the expression of 11 and 19 proteins, retene 29 and 23, while the mixture affected 44 and 82 DEPs by Days 7 and 14, respectively. In contrast, only 5 significantly affected metabolites were identified. Pathway over-representation analysis identified exposure-specific activation of phase II metabolic processes, which were accompanied with exposure-specific body burden profiles. The proteomic data highlights that exposure to the mixture increased oxidative stress, altered iron metabolism and impaired coagulation capacity. Additionally, depletion of several mini-chromosome maintenance components, in combination with depletion of several intermediate filaments and microtubules, among alevins exposed to the mixture, suggests compromised cellular integrity and reduced rate of mitosis, whereby affecting heart growth and development. Furthermore, the combination of proteomic and metabolomic data indicates altered energy metabolism, as per amino acid catabolism among mixture exposed alevins; plausibly compensatory mechanisms as to counteract reduced absorption and consumption of yolk. When considered as a whole, proteomic and metabolomic data, in relation to apical effects on the whole organism, provides additional insight into PAH toxicity and the effects of exposure on heart structure and molecular processes.</p

Admission criteria and management of critical care patients in a pandemic context: position of the Ethics Commission of the French Intensive Care Society, update of April 2021.

Intensive care unit professionals have experience in critical care and its proportionality, collegial decision-making, withholding or withdrawal of treatment deemed futile, and communication with patients' relatives. These elements rely on ethical values from which we must not deviate in a pandemic situation. The recommendations made by the Ethics Commission of the French Intensive Care Society reflect an approach of responsibility and solidarity towards our citizens regarding the potential impact of a pandemic on critical care resources in France, with the fundamental requirement of respect for human dignity and equal access to health care for all

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Utilisation d'un poisson modèle, Fundulus heteroclitus, pour évaluer le potentiel embryotoxique des polluants organiques persistants accumulés par l'anguille d'Amérique (Anguilla rostrata) du lac Ontario de 1988 à 2008

RÉSUMÉ: Le recrutement des juvéniles de l'anguille d'Amérique (Anguilla rostrata) vers le lac Ontario (LO), Canada, a décliné drastiquement depuis les années 1980. Afin d'explorer la contribution possible à ce déclin du transfert maternel des polluants organiques persistants (POP), la présente étude mesure les variations temporelles de la toxicité de mélanges organiques complexes extraits d'anguilles capturées dans le LO en 1988, 1998 et 2008, chez les embryons de Fundu/us heteroclitus exposés par injection intravitelline (IVi). Afin d'identifier les composés les plus toxiques accumulés par les anguilles et de caractériser leurs effets embryotoxiques, des embryons de F heteroclitus ont été exposés par injection IVi à des doses sous-létales de plusieurs POP historiquement mesurés dans les tissus des poissons du LO, incluant certains composés apparentés aux dioxines (TCDD, BPCI26, BPC77 et 2,3,4,7,8- PnCDF), des BPC non-coplanaires (BPC52 et BPCI10) ainsi qu'à un mélange technique de BPC, ]'Aroclor 1254. Tous les composés apparentés aux dioxines ainsi que l'Aroclor 1254 ont causé des malformations crânio-faciales, une réduction dose-dépendante de la croissance larvaire et de la capacité de prédation sans altération de l'activité locomotrice de base, ainsi qu'une induction de l'activité de l'ethoxyrésorufine-O-dééthylase (EROD) chez les larves de F heteroclitus. Aucune de ces réponses n'a été observée avec les BPC non-coplanaires. Les relations doseréponse pour l'activité EROD pour la TCDD, le BPCl26 et le 2,3,4,7,8-PnCDF avaient des pentes similaires et les potentiels toxiques relatifs (PTR) du BPC126 et du 2,3,4,7,8-PnCDF par rapport à la TCDD ont été estimés à 0.71 et 2.40, respectivement. Ils étaient respectivement environ 140 et 5 fois supérieurs aux facteurs d'équivalence toxique (FET) de référence de l'Organisation Mondiale de la Santé (OMS) pour ces deux composés chez les poissons, qui sont basés sur des données d' embryo-mortalité obtenues chez la truite arc-en-ciel (Oncorhynchus mykiss). Ces résultats suggèrent que les PTR sont spécifiques à chaque espèce et que les FET actuels du BPC126 et du 2,3,4,7,8-PnCDF pourraient sous-estimer leurs potentiels toxiques chez certaines espèces de poissons. Les pentes des relations dose-réponse pour la capacité de prédation étaient plus prononcées pour le BPC126 et le 2,3,4,7,8-PnCDF par rapport à la TCDD, suggérant différentes voies d'effets. Le niveau d'expression de plusieurs gènes a également été mesuré par réaction en chaîne par polymérase quantitative (qPCR) à la suite d'expositions à des doses uniques de TCDD ou de BPC126 (1280 et 1250 pg g-J de poids humide, respectivement) causant un même niveau d'induction EROD. Un patron de réponses différent a été observé entre le BPCl26 et la TCDD : le BPC126 a semblé induire des réponses anti-oxydantes par l'intermédiaire du gène sod2, mais pas la TCDD. Cela suggère que ces deux composés agissent selon des mécanismes différents pour induire les altérations comportementales observées chez les larves de F heteroclitus, et que le stress oxydant est un facteur clef dans le cas du BPCI26. Les extraits d'anguille de 1988 et 1998 ont été les plus toxiques, provoquant un patron de réponses embryotoxiques sous-létales similaire à celui observé chez des embryons de F heteroclitus exposés aux composés apparentés aux dioxines précédemment cités ou à l'Aroclor 1254 : retard de croissance, malformations crânio-faciales, induction de l'activité EROD et réduction de la capacité de prédation des larves. Le potentiel toxique des extraits a décliné avec le temps: le seul effet sign ificatif observé avec les extraits de 2008 était l'induction de l' activité EROD. Les autres POP mesurés dans les extraits d' anguilles (BPC non-coplanaires, PBDE et pesticides organochlorés) n'ont pas semblé être d' importants contributeurs à l'embryotoxicité observée. En raison de la combinaison de ses concentrations élevées et de son PTR élevé par rapport à la TCDD chez les jeunes stades de vie de F heteroclitus, le BPC 126 était probablement le plus important contributeur à l'embryotoxicité observée avec l'Aroclor 1254 et les extraits d' anguille. Néanmoins, comme cela a pu être observé avec l'Aroclor 1254, les EQT-TCDD dérivés des données chimiques, calculés à partir des concentrations de certains composés apparentés aux dioxines dans les extraits et de leurs PTR respectifs chez F heteroclitus, ont surestimé la capacité des extraits à induire l'activité EROD, possiblement en raison d' interactions entre les différents POP. La toxicité des mélanges organiques complexes de POP accumulés par les anguilles du LO a pu être sous-estimée en raison de plusieurs facteurs , incluant les pertes chimiques durant la préparation des extraits, ainsi que l'absence d'évaluation de leurs effets à long terme. En plus de différences interspécifiques en termes de sensibilité à la TCDD déjà documentées dans la littérature, nos résultats démontrent qu' il existe des différences interspécifiques de FEI. Il existe donc une incertitude quant à la prédiction de la toxicitié d' un mélange complexe de composés apparentés aux dioxines chez A. rostrata à partir de données obtenues chez d' autres espèces. Dans l'ensemble, nos résultats supportent l'hypothèse selon laquelle la contamination du LO par les composés apparentés aux dioxines aurait représenté une menace pour la population de l'anguille d'Amérique en altérant des paramètres pertinents d' un point de vue environnemental, telle que la capacité de prédation des larves. Ces résultats soulignent l'importance de tester l'embryotoxicité des composés apparentés aux dioxines chez les jeunes stades de vie des différentes espèces d' anguilles, d' explorer les effets à long terme d' extraits organiques d'anguilles sur les jeunes stades de vie des poissons et de développer des biomarqueurs pour évaluer les altérations potentielles chez les jeunes anguilles collectées sur le terrain. -- Mots-clés: Anguille d'Amérique, Fundulus heteroclitus, TCDD, dioxine, BPC, Aroclor, embryotoxicité, comportement, TEF, EROD. -- ABSTRACT: Recruitment of American eel (Anguilla rostrata) juveniles to Lake Ontario (LO), Canada, declined significantly since the 1980s. To investigate the possible contribution of maternally transferred persistent organic pollutants (POPs) to this deeline, this study measured temporal variations in the toxicity of complex organic mixtures extracted from LO eels captured in 1988, 1998 and 2008 to Fundulus heteroclitus embryos exposed by intravitelline (IVi) injection. To identify the most toxic compounds accumulated by eels and to characterize their embryotoxic effects, F heteroclitus embryos were exposed by IVi injection to sublethal doses of various POPs historically measured in LO fish tissues, including dioxinlike compounds (DLCs) (TCDD, PCB126, PCB77 and 2,3 ,4,7,8-PnCDF), non-dioxin-like (NDL) PCBs (PCB52 and PCB 110) and also to a complex mixture of PCBs, Aroelor 1254. All DLCs as weil as Aroelor 1254 caused craniofacial deformities, a dose-responsive reduction of larval growth and prey capture ability without alteration of basallocomotor activit y, and induction of ethoxyresorufin-O-deethylase (EROD) activity in F heteroclitus larvae. None of these responses was induced by the two NDL PCBs. The dose-response relationships for EROD activity for PCB 126, 2,3,4,7,8-PnCDF and TCDD had similar slopes and the relative potencies (RePs) of PCB126 and 2,3,4,7,8-PnCDF to TCDD for EROD activity were estimated at 0.71 and 2.40, respectively. They were approximately and respectively 140- and 5-fold higher than the WorId Health Organization (WHO) TCDD equivalency factors (TEFs) of PCB126 and 2,3,4,7,8-PnCDF for fish , which are based on rainbow trout (Oncorhynchus mykiss) embryolethality data. These results suggest that relative potencies are species-specific and that the current ReP for PCB126 and 2,3,4,7,8-PnCDF underestimates their toxicity for some fish species. The si opes of the dose-response relationships for prey capture ability for PCB126 and 2,3,4,7,8-PnCDF were steeper th an for TCDD, suggesting different pathways of effects. Expression levels of several genes were also studied by quantitative real-time polymerase chain reaction (qPCR) following exposure to single doses of TCDD or PCB 126 (1280 and 1250 pg g-I wet weight, respectively) causing similar EROD induction. A different pattern of responses was observed between PCB 126 and TCDD : PCB 126 appeared to induce antioxidant responses by inducing sod2 expression, while TCDD did not. It suggests that these two compounds act through different mechanisms to produce behavioral alterations in F heteroclitus larvae, and that oxidative stress may be a key factor in the case of PCB 126. The 1988 and 1998 eel ex tracts were the most toxic, causing a pattern of sublethal embryotoxic responses similar to those reported in F heteroclitus embryos exposed to previously cited DLCs or to Aroelor 1254 : stunted growth, craniofacial deformities, induction of EROD activity, and reduced predatory capacities of larvae. The potency of these extracts declined over time : the only significant effect of 2008 eel ex tracts was EROD induction . Other POPs measured in eel extracts (NDL PCBs, PBDEs and organochlorinated pesticides) did not appear to be important agonistic contributors to the observed toxicity. Due to the combination of its concentrations and its high ReP compared to TCDD in F heteroclitus early life stages (ELS), PCB 126 was probably the most important contributor to the ove ra li embryotoxicity observed with Aroelor 1254 or eel extracts. However, as observed with Aroelor 1254, the chemically derived TCDD-TEQs of eel extracts, calculated using measured concentrations of sorne DLCs and their RePs for F heteroclitus ELS, overestimated their potency to induce EROD activity possibly due to interactions among POPs. The toxicity of the complex mixtures of POPs accumulated by LO eels may have been underestimated as a result of several factors, ineluding the loss of POPs during ex tracts preparation and the non-assessment of their potentiallong-term effects. In addition to the interspecific differences in tenns of sensitivity to TCDD documented in the literature, our results demonstrates that interspecific differences also exist for TEFs. Thus, there is uncertainty in the prediction of the toxicity of complex mixtures of DLCs to A. rostrata from data obtained with other species. Overall, our results support the hypothesis that contamination of LO with DLCs may have represented a threat for the American eel population through ecologically relevant effects such as altered larval prey capture ability. These results prioritize the need to assess ELS toxicity of DLCs in Anguilla species, to investigate long-term or delayed effects of complex eel ex tracts to ELS of fish and to further develop biomarkers to assess potential effects in eel ELS col1ected in the field. -- Keywords : American eel, Fundulus heteroclitus, TCDD, dioxin, PCB, Aroclor, embryotoxicity, behavior, TEF, EROD

Salmo trutta is more sensitive than Oncorhynchus mykiss to early-life stage exposure to retene

Salmonids are known to be among the most sensitive fish to dioxin-like compounds (DLCs), but very little is known about the sensitivity of the brown trout (Salmo trutta), which has declined and is endangered in several countries of Europe and Western Asia. We investigated the sensitivity of brown trout larvae to a widespread dioxin-like PAH, retene (3.2 to 320 μg.L−1), compared to the larvae of a salmonid commonly used in toxicology studies, the rainbow trout (Oncorhynchus mykiss). Mortality, growth, cyp1a induction and the occurrence of deformities were measured after 15 days of exposure. Brown trout larvae showed a significantly higher mortality at 320 μg.L−1 compared to rainbow trout larvae. While the occurrence of deformities was only significantly increased at 320 μg.L−1 for the rainbow trout, brown trout larvae displayed pericardial edemas and hemorrhages already at 10 or 100 μg.L−1. cyp1a induction was increased significantly already at ≥3.2 μg.L−1 for the brown trout, versus ≥32 μg.L−1 for the rainbow trout. Least square regression analysis of the concentration-response relationships suggested that S. trutta larvae were at least 2 times more sensitive than O. mykiss larvae for cyp1a induction. The present study suggests that S. trutta larvae are more sensitive than O. mykiss larvae to a potent DLC, retene. As it is possible that S. trutta populations have declined partly because of pollution by DLCs, we recommend generating more data regarding the sensitivity of threatened fish populations, in order to ensure better risk assessment.peerReviewe

Sequencing dataset related to the influence of terrestrial plant litter input on microbial community composition and biomass in lake water

We compared decomposition processes of three deciduous tree leaves (Alnus sp., Betula sp., and Populus tremula) in lake water together and without lake water microbiome, which was killed by autoclaving to separately examine the microbial community succession and decomposition process without the effect of lake water microbiome. We followed microbial activity and community succession during the first three weeks and quantified microbial biomass and the proportion of fungal versus bacterial decomposers by sterol and phospholipid fatty acid (PLFA) analyses. The present dataset contains the raw sequencing files that were used to identify the microbial and fungal communities of the samples collected during the study

Exposure to retene, fluoranthene, and their binary mixture causes distinct transcriptomic and apical outcomes in rainbow trout (Oncorhynchus mykiss) yolk sac alevins

Polycyclic aromatic hydrocarbons (PAHs) are widely spread environmental contaminants which affect developing organisms. It is known that improper activation of the aryl hydrocarbon receptor (AhR) by some PAHs contributes to toxicity, while other PAHs can disrupt cellular membrane function. The exact downstream mechanisms of AhR activation remain unresolved, especially with regard to cardiotoxicity. By exposing newly hatched rainbow trout alevins (Oncorhynchus mykiss) semi-statically to retene (32 µg l−1; AhR agonist), fluoranthene (50 µg l−1; weak AhR agonist and CYP1a inhibitor) and their binary mixture for 1, 3, 7 and 14 days, we aimed to uncover novel mechanisms of cardiotoxicity using a targeted microarray approach. At the end of the exposure, standard length, yolk area, blue sac disease (BSD) index and PAH body burden were measured, while the hearts were prepared for microarray analysis. Each exposure produced a unique toxicity profile. We observed that retene and the mixture, but not fluoranthene, significantly reduced growth by Day 14 compared to the control, while exposure to the mixture increased the BSD-index significantly from Day 3 onward. Body burden profiles were PAH-specific and correlated well with the exposure-specific upregulations of genes encoding for phase I and II enzymes. Exposure to the mixture over-represented pathways related to growth, amino acid and xenobiotic metabolism and oxidative stress responses. Alevins exposed to the individual PAHs displayed over-represented pathways involved in receptor signaling: retene downregulated genes with a role in G-protein signaling, while fluoranthene upregulated those involved in GABA signaling. Furthermore, exposure to retene and fluoranthene altered the expression of genes encoding for proteins involved in calcium- and potassium ion channels, which suggests affected heart structure and function. This study provides deeper understanding of the complexity of PAH toxicity and the necessity of investigating PAHs as mixtures and not as individual components.peerReviewe

Metabolic plasticity of mixotrophic algae is key for their persistence in browning environments

Light availability is the main regulator of primary production, shaping photosynthetic communities and their production of ecologically important biomolecules. In freshwater ecosystems, increasing dissolved organic carbon concentrations, commonly known as browning, leads to lower light availability and the proliferation of mixotrophic phytoplankton. Here, a mixotrophic algal species (Cryptomonas sp.) was grown under five increasing dissolved organic carbon concentrations to uncover the plastic responses behind the success of mixotrophs in browning environments and their effect in the availability of nutritionally important biomolecules. In addition to the browning treatments, phototrophic, heterotrophic and mixotrophic growth conditions were used as controls. Despite reduced light availability, browning did not impair algal growth compared to phototrophic conditions. Comparative transcriptomics showed that genes related to photosynthesis were down-regulated, whereas phagotrophy gene categories (phagosome, lysosome, and endocytosis) were up-regulated along the browning gradient. Stable isotope analysis of phospholipid fractions validated these results, highlighting that the studied mixotroph increases its reliance on heterotrophic processes with browning. Metabolic pathway reconstruction using transcriptomic data suggests that organic carbon is acquired through phagotrophy and used to provide energy in conjunction with photosynthesis. Although metabolic responses to browning were observed, essential fatty acid content was similar between treatments while sterol content was slightly higher upon browning. Altogether, our results provide a mechanistic model of how a mixotrophic algae responds to browning and how such responses affect the availability of nutritionally essential biomolecules for higher trophic levels.peerReviewe

Relative potency of PCB126 to TCDD for sublethal embryotoxicity in the mummichog (Fundulus heteroclitus)

International audienceThe relative potency (ReP) of 3,3',4,4',5-pentachlorobiphenyl (PCB126) to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for sublethal responses was assessed in Fundulus heteroclitus embryos. Eggs were treated with intravitelline injections of graded sublethal doses of PCB126 (312-5000 pg g(-1) wet weight, ww) or TCDD (5-1280 pg g(-1) ww). At 16 days post-fertilization (DPF), craniofacial deformities were observed in larvae hatched from eggs treated with the two highest doses of PCB126 (2500-5000 pg g(-1) ww). Both compounds caused a dose-responsive reduction of larval growth and prey capture ability (at ≥1250 pg g(-1) ww), and induction of ethoxyresorufin-O-deethylase (EROD) activity (at ≥80 pg g(-1) ww). The dose-response relationships for EROD activity for PCB126 and TCDD had similar slopes and the ReP of PCB126 to TCDD for EROD activity was estimated at 0.71. This is 140-fold higher than the World Health Organization (WHO) TCDD equivalency factor (TEF) of PCB126 for fish (0.005), which is based on rainbow trout (Oncorhynchus mykiss) embryolethality data. The slope of the dose-response relationship for prey capture ability for PCB126 was steeper than for TCDD, suggesting different mechanisms of action. Expression levels of several genes were also studied by quantitative real-time polymerase chain reaction (qPCR) following exposure to single doses of TCDD or PCB126 (1280 and 1250 pg g(-1) ww, respectively) causing similar EROD induction. A different pattern of responses was observed between PCB126 and TCDD: PCB126 appeared to induce antioxidant responses by inducing sod2 expression, while TCDD did not. These results suggest that relative potencies are species-specific and that the current ReP for PCB126 underestimates its toxicity for some fish species. It is recommended to develop species-specific RePs for a variety of sublethal endpoints and at environmentally relevant doses