How the Xenopus eleutheroembryonic thyroid assay compares to the amphibian metamorphosis assay for detecting thyroid active chemicals.

The Xenopus Eleutheroembryonic Thyroid Assay (XETA) was recently published as an OECD Test Guideline for detecting chemicals acting on the thyroid axis. However, the OECD validation did not cover all mechanisms that can potentially be detected by the XETA. This study was therefore initiated to investigate and consolidate the applicability domain of the XETA regarding the following mechanisms: thyroid hormone receptor (THR) agonism, sodium-iodide symporter (NIS) inhibition, thyroperoxidase (TPO) inhibition, deiodinase (DIO) inhibition, glucocorticoid receptor (GR) agonism, and uridine 5'-diphospho-glucuronosyltransferase (UDPGT) induction. In total, 22 chemicals identified as thyroid-active or -inactive in Amphibian Metamorphosis Assays (AMAs) were tested using the XETA OECD Test Guideline. The comparison showed that both assays are highly concordant in identifying chemicals with mechanisms of action related to THR agonism, DIO inhibition, and GR agonism. They also consistently identified the UDPGT inducers as thyroid inactive. NIS inhibition, investigated using sodium perchlorate, was not detected in the XETA. TPO inhibition requires further mechanistic investigations as the reference chemicals tested resulted in opposing response directions in the XETA and AMA. This study contributes refining the applicability domain of the XETA, thereby helping to clarify the conditions where it can be used as an ethical alternative to the AMA.

Insect-based fish feed in decoupled aquaponic systems: Effect on lettuce production and resource use.

The utilisation of insect meal-based fish feed as a substitute for conventional fish meal-based fish feed is considered as a promising innovative alternative to boost circularity in aquaculture and aquaponics. Basic research on its use in aquaponics is limited. So far, no reports on the effects of fish waste water, derived from a recirculating aquaculture system using Black Soldier Fly (BSF) meal-based diets, were available on the growth performance of lettuce. Therefore, this study aimed to compare the effect of reusing fish waste water from tilapia culture (as a base for the nutrient solution) fed with a fish meal-based diet (FM) and a BSF meal-based diet on resource use and lettuce growth in decoupled aquaponic systems. A conventional hydroponics nutrient solution (HP) served as control, and inorganic fertilisers were added to all nutrient solutions to reach comparable target concentrations. The experiment was conducted in a controlled climate chamber in nine separate hydroponics units, three per treatment. Lettuce fresh and dry weight, number of leaves, relative leaf chlorophyll concentration, water consumption, and the usage of inorganic fertilisers were measured. Micro- and macronutrients in the nutrient solutions were monitored in time series. Similar lettuce yield was seen in all treatments, with no significant effects on fresh and dry weight, the number of leaves, and relative chlorophyll values. Water use per plant was also similar between treatments, while the amount of total inorganic fertiliser required was 32% lower in FM and BSF compared to HP. Higher sodium concentrations were found in the FM nutrient solutions compared to BSF and HP. The results confirm that BSF-based diet is a promising alternative to FM-based diet in aquaponics with no negative effects on lettuce growth. Additionally, BSF-based diet might be beneficial in intensive, professional aquaponics applications due to the lower sodium concentration in the nutrient solution.

Detection of anti-androgenic activity of chemicals in fish studies: a data review.

A systematic review was conducted on the sensitivity of fish testing guidelines to detect the anti-androgenic activity of substances. Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) was used to investigate the conservation of the androgen receptor (AR) between humans and fish, and among fish species recommended in test guidelines. The AR is conserved between fish species and humans (i.e. ligand binding domain [LBD] homology ≥70%) and among the recommended fish species (LBD homology >85%). For model anti-androgens, we evaluated literature data on in vitro anti-androgenic activity in fish-specific receptor-based assays and changes in endpoints indicative of endocrine modulation from in vivo studies. Anti-androgenic activity was most consistently and reliably detected in in vitro and in vivo mechanistic studies with co-exposure to an androgen (spiggin in vitro assay, Rapid Androgen Disruption Activity Reporter [RADAR] Assay, and Androgenised Female Stickleback Screen). Regardless of study design (Fish Short-Term Reproduction Assay [FSTRA], Fish Sexual Development Test [FSDT], partial or full life-cycle tests), or endpoint (vitellogenin, secondary sexual characteristics, gonadal histopathology, sex ratio), there was no consistent evidence for detecting anti-androgenic activity in studies without androgen co-exposure, even for the most potent substances (while less potent substances may induce no (clear) response). Therefore, based on studies without androgen co-exposure (35 FSTRAs and 22 other studies), the other studies (including the FSDT) do not outperform the FSTRA for detecting potent anti-androgenic activity, which if suspected, would be best addressed with a RADAR assay. Overall, fish do not appear particularly sensitive to mammalian anti-androgens.

Make a choice: A rapid strategy for minimizing peat in horticultural press pots substrates using a constrained mixture design and surface response approach.

Peat is the most common used substrate in horticultural seedling production. To reduce peat in horticultural potted plant cultivation systems in general is an obstacle, even within the highly specialized horticultural industry. Next to soil-less cultivation systems as e.g. hydroponics, the horticultural industry is eagerly looking for suitable peat substitutes. The demands on these compounds are high, basically mimicking the physical properties of peat. A 100% replacement of peat for press-pots used in seedling production has not yet been found, and only mixes of peat and substrates exist. Several suitable peat substitutes with different properties are known, that usually are used as a share of a mixed peat-substitute substrate. A constrained mixture design was used to test substrates containing 50% v/v and 25% v/v peat and four peat substitutes (two composts and two wood fibers) for vegetable seedling production. By limiting the maximum quantities of each material to be added, there was no negative effect on the growth of Chinese cabbage (Brassica rapa subsp. pekinensis). This means a reduction in of peat to 25% v/v is possible without a change in substrate quality. The mixture design allowed a quick decision to be made regarding the most suitable peat-reduced mixtures. The surface response approach enabled the experimental results to be easily transferred to horticultural practices, additionally. This flexible and efficient method also allows the predictions to be used to meet specific crop management needs.

Using leaf spectroscopy and pigment estimation to monitor indoor grown lettuce dynamic response to spectral light intensity.

Rising urban food demand is being addressed by plant factories, which aim at producing quality food in closed environment with optimised use of resources. The efficiency of these new plant production systems could be further increased by automated control of plant health and nutritious composition during cultivation, allowing for increased produce value and closer match between plant needs and treatment application with potential energy savings. We hypothesise that certain leaf pigments, including chlorophylls, carotenoids and anthocyanins, which are responsive to light, may be good indicator of plant performance and related healthy compounds composition and, that the combination of leaf spectroscopy and mathematical modelling will allow monitoring of plant cultivation through noninvasive estimation of leaf pigments. Plants of two lettuce cultivars (a green- and a red-leaf) were cultivated in hydroponic conditions for 18 days under white light spectrum in climate controlled growth chamber. After that period, plant responses to white light spectrum ('W') with differing blue wavelengths ('B', 420 - 450 nm) percentage (15% 'B15', and 40% 'B40') were investigated for a 14 days period. The two light spectral treatments were applied at photon flux densities (PFDs) of 160 and 240 µmol m-2 s-1, resulting in a total of four light treatments (160WB15, 160WB40, 240WB15, 240WB40). Chlorophyll a fluorescence measurements and assessment of foliar pigments, through destructive (in vitro) and non-destructive (in vivo) spectrophotometry, were performed at 1, 7 and 14 days after treatment initiation. Increase in measured and estimated pigments in response to WB40 and decrease in chlorophyll:carotenoid ratio in response to higher PFD were found in both cultivars. Cultivar specific behavior in terms of specific pigment content stimulation in response to time was observed. Content ranges of modelled and measured pigments were comparable, though the correlation between both needs to be improved. In conclusion, leaf pigment estimation may represent a potential noninvasive and real-time technique to monitor, and control, plant growth and nutritious quality in controlled environment agriculture.

Response of Cyanic and Acyanic Lettuce Cultivars to an Increased Proportion of Blue Light.

Indoor crop cultivation systems such as vertical farms or plant factories necessitate artificial lighting. Light spectral quality can affect plant growth and metabolism and, consequently, the amount of biomass produced and the value of the produce. Conflicting results on the effects of the light spectrum in different plant species and cultivars make it critical to implement a singular lighting solution. In this study we investigated the response of cyanic and acyanic lettuce cultivars to an increased proportion of blue light. For that, we selected a green and a red leaf lettuce cultivar (i.e., 'Aquino', CVg, and 'Barlach', CVr, respectively). The response of both cultivars to long-term blue-enriched light application compared to a white spectrum was analyzed. Plants were grown for 30 days in a growth chamber with optimal environmental conditions (temperature: 20 °C, relative humidity: 60%, ambient CO2, photon flux density (PFD) of 260 µmol m-2 s-1 over an 18 h photoperiod). At 15 days after sowing (DAS), white spectrum LEDs (WW) were compared to blue-enriched light (WB; λPeak = 423 nm) maintaining the same PFD of 260 µmol m-2 s-1. At 30 DAS, both lettuce cultivars adapted to the blue light variant, though the adaptive response was specific to the variety. The rosette weight, light use efficiency, and maximum operating efficiency of PSII photochemistry in the light, Fv/Fm', were comparable between the two light treatments. A significant light quality effect was detected on stomatal density and conductance (20% and 17% increase under WB, respectively, in CVg) and on the modified anthocyanin reflectance index (mARI) (40% increase under WB, in CVr). Net photosynthesis response was generally stronger in CVg compared to CVr; e.g., net photosynthetic rate, Pn, at 1000 µmol m-2 s-1 PPFD increased from WW to WB by 23% in CVg, compared to 18% in CVr. The results obtained suggest the occurrence of distinct physiological adaptive strategies in green and red pigmented lettuce cultivars to adapt to the higher proportion of blue light environment.

A Photosynthetic Light Acclimation Model Accounting for the Effects of Leaf Age, Chlorophyll Content, and Intra-Leaf Radiation Transfer.

Mechanistic models of canopy photosynthesis usually upscale leaf photosynthesis to crop level. A detailed prediction of canopy microclimate with accurate leaf morphological and physiological model parameters is the pre-requisite for accurate predictions. It is well established that certain leaf model parameters (V cmax, J max) of the frequently adopted Farquhar and Caemmerer photosynthesis model change with leaf age and light interception history. Previous approaches to predict V cmax and J max focused primarily on light interception, either by cumulative intercepted photosynthetic photon flux density (PPFD) or by closely related proxy variables such as leaf nitrogen content per leaf area. However, for plants with monopodial growth, such as vertically grown tomatoes or cucumber crops, in greenhouse production, there is a strong relationship between leaf age and light interception, complicating the experimental and mathematical separation of both effects. We propose a modeling framework that separates age and light intensity-related acclimation effects in a crop stand: Improved approximation of intra-leaf light absorption profiles with cumulative chlorophyll content (Chl) is the basis, while parameters are estimated via Gaussian process regression from total Chl, carotenoid content (Car), and leaf mass per area (LMA). The model approximates light absorption profiles within a leaf and links them to leaf capacity profiles of photosynthetic electron transport. Published datasets for Spinacia oleracea and Eucalyptus pauciflora were used to parameterize the relationship between light and capacity profiles and to set the curvature parameter of electron transport rate described by a non-rectangular hyperbola on Cucumis sativus. Using the modified capacity and light absorption profile functions, the new model was then able to predict light acclimation in a 2-month period of a fully grown tomato crop. An age-dependent lower limit of the electron transport capacity per unit Chl was essential in order to capture the decline of V cmax and J max over time and space of the investigated tomato crop. We detected that current leaf photosynthetic capacity in tomato is highly affected by intercepted light-sum of 3-5 previous days.

Crop Photosynthetic Performance Monitoring Based on a Combined System of Measured and Modelled Chloroplast Electron Transport Rate in Greenhouse Tomato.

Combining information of plant physiological processes with climate control systems can improve control accuracy in controlled environments as greenhouses and plant factories. Through that, resource optimization can be achieved. To predict the plant physiological processes and implement them in control actions of interest, a reliable monitoring system and a capable control system are needed. In this paper, we focused on the option to use real-time crop monitoring for precision climate control in greenhouses. For that, we studied the processes and external factors influencing leaf net CO2 assimilation rate (AL , µmol CO2 m-2 s-1) as possible variables of a plant performance indicator. While measured greenhouse environmental variables such as light, temperature, or humidity showed a direct relation between AL and light-quantum yield of photosystem II (Φ2), we defined three objectives: (1) to explore the relationship between climate variables and AL , as well as Φ2; (2) create a simple and reliable method for real-time prediction of AL with continuously Φ2 measurements; and (3) calibrate parameters to predict chloroplast electron transport rate as input in AL modelling. Due to practical obstacles in measuring CO2 gas-exchange in commercial production, we explored a method to predict AL by measuring Φ2 of leaves in a commercial hydroponic greenhouse tomato crop ("Pureza"). We calculated AL with two different approaches based on either the negative exponential response model with simplified biochemical equations (marked as Model I) or the non-rectangular hyperbola full biochemical photosynthetic models (marked as Model II). Using Model I can only be used to predict AL with large uncertainty (R2 0.64; RMSE 2.21), while using Φ2 as input to Model II could be used to improve the prediction accuracy of AL (R2 0.71; RMSE 1.98). Our results suggests that (1) Φ2 light signals can be used to predict net photosynthesis rate with high accuracy; (2) a parameterized photosynthetic electron transport rate model is suitable predicting measured electron transport rate (J) and AL . The system can be used as decision support system (DSS) for plant and crop performance monitoring when leaf-dynamics are up-scaled to the plant or crop level.

Stomatal behavior following mid- or long-term exposure to high relative air humidity: A review.

High relative air humidity (RH ≥ 85%) is frequent in controlled environments, and not uncommon in nature. In this review, we examine the high RH effects on plants with a special focus on stomatal characters. All aspects of stomatal physiology are attenuated by elevated RH during leaf expansion (long-term) in C3 species. These include impaired opening and closing response, as well as weak diel oscillations. Consequently, the high RH-grown plants are not only vulnerable to biotic and abiotic stress, but also undergo a deregulation between CO2 uptake and water loss. Stomatal behavior of a single leaf is determined by the local microclimate during expansion, and may be different than the remaining leaves of the same plant. No effect of high RH is apparent in C4 and CAM species, while the same is expected for species with hydropassive stomatal closure. Formation of bigger stomata with larger pores is a universal response to high RH during leaf expansion, whereas the effect on stomatal density appears to be species- and leaf side-specific. Compelling evidence suggests that ABA mediates the high RH-induced stomatal malfunction, as well as the stomatal size increase. Although high RH stimulates leaf ethylene evolution, it remains elusive whether or not this contributes to stomatal malfunction. Most species lose stomatal function following mid-term (4-7 d) exposure to high RH following leaf expansion. Consequently, the regulatory role of ambient humidity on stomatal functionality is not limited to the period of leaf expansion, but holds throughout the leaf life span.

Online measurement of temperature and relative humidity as marker tools for quality changes in onion bulbs during storage.

A long shelf life of onions (Allium cepa L.) is of high importance in the onion industry. Onions are dried and stored in large wooden boxes that are difficult to access. Monitoring temperature and relative humidity during these processes is challenging. Moreover, quality may change in storage without being noticed. Therefore, there is a need to find alternative methods for monitoring and controlling the drying and storage processes of onions and to identify early changes in quality during storage. The potential use of online measurements of temperature and relative humidity (RH) in the vicinity of onions was evaluated during drying and long-term storage of six onion batches (four cultivars and three selections of one of the cultivars) in commercial storage. The batches varied in bulb weight, dry matter content, firmness and disease incidence. The dry matter content and firmness decreased during storage, while the respiration rate and incidences of individual and total disease increased. Two of the batches had low storability with high disease incidences and high average temperatures and variations in the RH. The results showed that tracking the temperature and RH in the vicinity of the onions is a promising tool for improving the drying and storage processes in commercial storage and for identifying onion batches with reduced storability early in storage.

Using the quantum yields of photosystem II and the rate of net photosynthesis to monitor high irradiance and temperature stress in chrysanthemum (Dendranthema grandiflora).

Under a dynamic greenhouse climate control regime, temperature is adjusted to optimise plant physiological responses to prevailing irradiance levels; thus, both temperature and irradiance are used by the plant to maximise the rate of photosynthesis, assuming other factors are not limiting. The control regime may be optimised by monitoring plant responses, and may be promptly adjusted when plant performance is affected by extreme microclimatic conditions, such as high irradiance or temperature. To determine the stress indicators of plants based on their physiological responses, net photosynthesis (Pn) and four chlorophyll-a fluorescence parameters: maximum photochemical efficiency of PSII [Fv/Fm], electron transport rate [ETR], PSII operating efficiency [F'q/F'm], and non-photochemical quenching [NPQ] were assessed for potted chrysanthemum (Dendranthema grandiflora Tzvelev) 'Coral Charm' under different temperature (20, 24, 28, 32, 36 °C) and daily light integrals (DLI; 11, 20, 31, and 43 mol m(-2) created by a PAR of 171, 311, 485 and 667 µmol m(-2) s(-1) for 16 h). High irradiance (667 µmol m(-2) s(-1)) combined with high temperature (>32 °C) significantly (p < 0.05) decreased Fv/Fm. Under high irradiance, the maximum Pn and ETR were reached at 24 °C. Increased irradiance decreased the PSII operating efficiency and increased NPQ, while both high irradiance and temperature had a significant effect on the PSII operating efficiency at temperatures >28 °C. Under high irradiance and temperature, changes in the NPQ determined the PSII operating efficiency, with no major change in the fraction of open PSII centres (qL) (indicating a QA redox state). We conclude that 1) chrysanthemum plants cope with excess irradiance by non-radiative dissipation or a reversible stress response, with the effect on the Pn and quantum yield of PSII remaining low until the temperature reaches 28 °C and 2) the integration of online measurements to monitor photosynthesis and PSII operating efficiency may be used to optimise dynamic greenhouse control regimes by detecting plant stress caused by extreme microclimatic conditions.

High temperature stress monitoring and detection using chlorophyll a fluorescence and infrared thermography in chrysanthemum (Dendranthema grandiflora).

Modern highly insulated greenhouses are more energy efficient than conventional types. Furthermore applying dynamic greenhouse climate control regimes will increase energy efficiency relatively more in modern structures. However, this combination may result in higher air and crop temperatures. Too high temperature affects the plant photosynthetic responses, resulting in a lower rate of photosynthesis. To predict and analyse physiological responses as stress indicators, two independent experiments were conducted, to detect the effect of high temperature on photosynthesis: analysing photosystem II (PSII) and stomatal conductance (gs). A combination of chlorophyll a fluorescence, gas exchange measurements and infrared thermography was applied using Chrysanthemum (Dendranthema grandiflora Tzvelev) 'Coral Charm' as a model species. Increasing temperature had a highly significant effect on PSII when the temperature exceeded 38 °C for a period of 7 (±1.8) days. High temperature decreased the maximum photochemical efficiency of PSII (Fv/Fm), the conformation term for primary photochemistry (Fv/Fo) and performance index (PI), as well as increased minimal fluorescence (Fo). However, at elevated CO2 of 1000 µmol mol(-1) and with a photosynthetic photon flux density (PPFD) of 800 µmol m(-2) s(-1), net photosynthesis (Pn) reached its maximum at 35 °C. The thermal index (IG), calculated from the leaf temperature and the temperature of a dry and wet reference leaf, showed a strong correlation with gs. We conclude that 1) chlorophyll a fluorescence and a combination of fluorescence parameters can be used as early stress indicators as well as to detect the temperature limit of PSII damage, and 2) the strong relation between gs and IG enables gs to be estimated non-invasively, which is an important first step in modelling leaf temperature to predict unfavourable growing conditions in a (dynamic) semi closed greenhouse.

Water temperature and concomitant waterborne ethinylestradiol exposure affects the vitellogenin expression in juvenile brown trout (Salmo trutta).

Environmental estrogens have the potential to considerably affect the reproduction and development of aquatic vertebrates by interfering with the endocrine system. In addition to the potential risk of environmental estrogens, increasing water temperatures as a result of global warming have become a serious problem in many rivers and streams. To assess the degree of estrogenic exposure, the analysis of the estrogen-dependent protein vitellogenin (Vtg) is a frequently used biomarker in field studies. Little, however, is known regarding the potential interaction between ambient water temperature and the Vtg production induced by waterborne environmental estrogens. In order to test the influence of temperature on Vtg synthesis, we exposed juvenile brown trout to an environmentally relevant concentration of ethinylestradiol (EE(2)) and held them either at low or high temperatures (12 and 19 degrees C, respectively), but also at temperature cycles of 12-19 degrees C in order to simulate the field situation. The EE(2) exposure caused a 7-74-fold increase of hepatic Vtg mRNA. The synthesis of Vtg mRNA was clearly stimulated in fish held at higher water temperatures (12-19 degrees C and 19 degrees C, respectively). On the protein level, Vtg showed a similar pattern; the higher the temperature, the higher the concentration of Vtg in the plasma. The experiment further revealed a temperature-dependent increasing amount of hepatic estrogen receptor alpha mRNA (ERalpha) after exposure to waterborne EE(2). The gene expression of estrogen receptor beta-1 (ERbeta-1) and the glucocorticoid receptor (GR) in the liver of EE(2) exposed fish, however, showed no treatment-related alterations. In line with observed constant bile cortisol concentrations, our data do not indicate corresponding stress related effects on hepatic Vtg production. The present survey, however, clearly demonstrates that increased temperature significantly elevates the estrogen-induced expression of Vtg and therefore has to be considered when interpreting environmental monitoring studies.

Assessment of estrogenic exposure in brown trout (Salmo trutta) in a Swiss midland river: integrated analysis of passive samplers, wild and caged fish, and vitellogenin mRNA and protein.

This field study examined the vitellogenin (VTG) biomarker response under conditions of low and fluctuating activities of environmental estrogenicity. The present study was performed on immature brown trout (Salmo trutta) exposed to the small river Luetzelmurg, which is located in the prealpine Swiss midland region and receives effluents from a single sewage treatment plant (STP). To understand better factors influencing the relationship between estrogenic exposure and VTG induction, we compared VTG levels in caged (stationary) and feral (free-ranging) fish, VTG levels in fish from up- and downstream of the STP, and two different methods for quantifying VTG (enzyme-linked immunosorbent assay vs real-time reverse transcription-polymerase chain reaction), and we used passive samplers (polar organic chemical integrative sampler [POCIS]) to integrate the variable, bioaccumulative estrogenic load in the river water over time. The POCIS from the downstream site contained approximately 20-fold higher levels of bioassay-derived estrogen equivalents than the POCIS from the upstream site. In feral fish, this site difference in estrogenic exposure was reflected in VTG protein levels but not in VTG mRNA. In contrast, in caged fish, the site difference was evident only for VTG mRNA but not for VTG protein. Thus, the outcome of VTG biomarker measurements varied with the analytical detection method (protein vs mRNA) and with the exposure modus (caged vs feral). Our findings suggest that for environmental situations with low and variable estrogenic contamination, a multiple-assessment approach may be necessary for the assessment of estrogenic exposure in fish.

Characterization of environmental estrogens in river water using a three pronged approach: active and passive water sampling and the analysis of accumulated estrogens in the bile of caged fish.

Estrogenicity of river water is highly variable and it is difficult to obtain an average measure of the estrogenicity. Consequently it is difficult to tie the estrogenic effects observed in fish to their level of exposure to estrogens. To get a better handle on average estrogenic exposure we tested a recently developed passive sampling system (polar organic chemical integrative sampler, POCIS). In addition, we investigated the bioaccumulation of estrogens in caged brown trout and measured plasma vitellogenin in males as a bioindicator of estrogenic effects. We developed a mini-caging method to suit the hydrological conditions in small rivers and to improve upon the often poor survival of salmonids in caging trials. POCISs were positioned upstream and downstream of 5 sewage treatment works' discharges and left on site for 3 weeks (as were the caged fish), during which period 3 water grab samples were taken at each site. Concentrations of estrogens were determined using a yeast-based reporter gene assay and chemical analysis. Results from grab sampling, passive sampling, and bioaccumulation were correlated; however, plasma vitellogenin concentrations were elevated at only 1 of 5 sites. POCISs provide an integrated and biologically meaningful measure of estrogenicity in thatthey accumulate estrogens in a pattern similar to that of brown trout. Mini-caging appears a significant methodological advance; no fish were lost, moreover, all fish survived in excellent health.