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Mangrove forests thrive along tropical and subtropical shorelines and their distribution extends to warm temperate regions1. They are globally recognized for the valuable ecosystem services they provide2 but are expected to be substantially influenced by climate change-related physical processes in the future3,4. Under warming winter temperatures, poleward expansion is predicted for mangroves5,6, with potential implications for ecosystem structure and functioning, as well as human livelihoods and well-being7,8. The global distribution, abundance and species richness of mangroves is governed by a broad range of biotic and environmental factors, including temperature and precipitation9 and diverse geomorphological and hydrological gradients10. Climate and aspects related to coastal geography (for example, floodplain area) determine the availability of suitable habitat for establishment11,12. However, the potential for mangroves to track changing environmental conditions and expand their distributions ultimately depends on dispersal11,13. The importance of dispersal in controlling mangrove distributions has been demonstrated by mangrove distributional responses to historical climate variability14, past mangrove (re)colonization of oceanic islands15 and from the long-term survival of mangrove seedlings planted beyond natural range limits16. As such, quantifying changes in the factors that influence dispersal is important for understanding climate-driven distributional responses of mangroves under future climate conditions.In mangroves, dispersal is accomplished by buoyant seeds and fruits (hereafter referred to as ‘propagules’). In combination with prevailing currents, the spatial scale of this process, ranging from local retention to transoceanic dispersal over thousands of kilometres13, is determined by propagule buoyancy17, that is, the density difference between that of propagules and the surrounding water. Hence, the course of dispersal trajectories for propagules from these species depends on the interaction between spatiotemporal changes in both propagule density and that of the surrounding water, rendering this process sensitive to climate-driven changes in coastal and open-ocean water properties. The biogeographic implications of such density differences were recognized more than a century ago by Henry Brougham Guppy, who discussed18 ‘the far-reaching influence on plant-distribution and on plant-development that the relation between the specific weight of seeds and fruits and the density of sea-water must possess’.Since the time of Guppy’s early observations, climate change from human activities has driven pronounced changes in ocean temperature and salinity, with further changes predicted throughout the twenty-first century19. Ocean density is a nonlinear function of temperature, salinity and pressure20; therefore, these changes may influence dispersal patterns of mangrove propagules by altering their buoyancy and floating orientation. As Guppy noted18, ‘[for] plants whose seeds or fruits are not much lighter than seawater […] the effect of increased density of the water is to extend the flotation period’ or ‘to increase the number that floated for a given period’. Guppy also reported that the seedlings of the widespread mangrove genera Rhizophora and Bruguiera present exceptional examples of propagules with densities somewhere between seawater and freshwater18. Previous studies of the impacts of climate change on mangroves have focused on factors such as sea level rise, altered precipitation regimes and increasing temperature and storm frequency4,21,22,23 but the potential impact of climate-driven changes in seawater properties on mangroves has not yet been examined. This is somewhat surprising, as the ocean is the primary dispersal medium of this ‘sea-faring’ coastal vegetation and dispersal is a key process that governs a species’ response to climate change by changing its geographical range. This knowledge gap contrasts with recent efforts to expose links between climate change and dispersal in other ecologically important marine taxa such as zooplankton and fish species24,25,26,27.In this study, we investigate predicted changes in sea surface temperature (SST), sea surface salinity (SSS) and sea surface density (SSD) for coastal waters bordering mangrove forests (hereafter referred to as ‘coastal mangrove waters’), over the next century. Using a biogeographic classification system for coastal and shelf areas28, we examine spatiotemporal changes in these surface ocean properties, with a particular focus on the world’s two major mangrove diversity hotspots: (1) the Atlantic East Pacific (AEP) region, including all of the Americas, West and Central Africa and (2) the Indo West Pacific (IWP) region, extending from East Africa eastwards to the islands of the central Pacific1. Finally, we synthesize available data on the density of mangrove propagules for different mangrove species and explore the potential impact of climate-driven changes in SSD on propagule dispersal.To assess changes in SST and SSS throughout the global range of mangrove forests, we used present (2000–2014) and future (2090–2100) surface ocean properties from the Bio-ORACLE database29,30. SSD estimates were derived from these variables using the UNESCO EOS-80 equation of state polynomial for seawater31. Changes in SST, SSS and SSD (Fig. 1) were calculated for four representative concentration pathways (RCPs) and derived for coastal waters closest to the 583,578 polygon centroids from the 2015 Global Mangrove Watch (GMW) database32. After removing duplicates, our dataset contained 10,108 unique mangrove occurrence locations, with corresponding present conditions and predicted future changes in mean SST, SSS and SSD. Under the low-warming scenario RCP 2.6, mean SST of coastal mangrove waters is predicted to change by +0.64 (±0.11) °C and mean SSS by −0.06 (±0.25) practical salinity units (PSU). Combined, this results in an average change in mean SSD of −0.25 (±0.20) kg m−3 in coastal mangrove waters by the late twenty-first century (Supplementary Table 1). These values roughly double under RCP 4.5 (Supplementary Table 2), while under RCP 6.0, a change of +1.69 (±0.14) °C in mean SST, −0.21 (±0.42) PSU in mean SSS and −0.71 (±0.32) kg m−3 in mean SSD is predicted (Supplementary Table 3). Under RCP 8.5, our study predicts a change in SST of +2.84 (±0.21) °C (range 2.11–4.01 °C), a change in SSS of −0.30 (±0.74) PSU (−2.01–1.26 PSU) and a corresponding change in SSD of −1.17 (±0.56) kg m−3 (−2.53–0.03 kg m−3) (Supplementary Table 4).Fig. 1: Global map showing the change in sea surface variables across mangrove bioregions under RCP 8.5.a–c, Change in SST (a), SSS (b) and SSD (c). Changes in SST and SSS are based on present-day (2000–2014) and future (2090–2100) marine fields from the Bio-ORACLE database29,30, from which SSD data were derived. The vertical line (19° E) separates the two major mangrove bioregions: the AEP and IWP.Full size imageSpatial variability in predicted surface ocean property changes was examined by considering the two major mangrove bioregions (AEP and IWP) (Fig. 2) and using the Marine Ecoregions of the World (MEOW) biogeographic classification28 (Fig. 3). Both the range and changes in mean SST were comparable for the AEP and IWP mangrove bioregions, for all respective RCP scenarios (Fig. 2a and Supplementary Tables 1–4). Under RCP 8.5, mean SST in both mangrove bioregions is predicted to warm ~2.8 °C by 2100, which is roughly 4.5 times the predicted increase in mean SST under RCP 2.6 (Supplementary Tables 1 and 4). Predictions for the RCP 8.5 scenario are generally consistent with reported global ocean temperature trends33 and show that the greatest warming occurs in coastal waters near the Galapagos Islands (change in mean SST of 3.92 ± 0.06 °C). Pronounced SST increases are also predicted for Hawaii (change in mean SST of 3.36 ± 0.05 °C), the Southeast Australian Shelf (3.30 ± 0.25 °C), Northern and Southern New Zealand (3.25 ± 0.07 °C and 3.34 ± 0.02 °C, respectively), Warm Temperate Northwest Pacific (3.27 ± 0.16 °C), the Red Sea and Gulf of Aden (3.24 ± 0.08 °C), Somali/Arabian Coast (3.23 ± 0.15 °C), South China Sea (3.07 ± 0.10 °C), the Tropical East Pacific (3.09 ± 0.15 °C) and the Warm Temperate Northwest Atlantic (3.14 ± 0.13 °C) (Fig. 3b and Supplementary Tables 4).Fig. 2: Change in surface ocean properties for coastal waters bordering mangrove forests and in the two major mangrove bioregions, the AEP and IWP, for different RCPs.a–c, Variation in SST (a), SSS (b) and SSD (c) under various RCP scenarios. Grey indicates global distribution (n = 10,108), orange denotes AEP (n = 3,190) and green represents IWP (n = 6,918). Data for SST and SSS consist of present-day (2000–2014) and future (2090–2100) marine fields from the Bio-ORACLE database29,30, from which SSD data were derived. The cat-eye plots50 show the distribution of the data. Median and mean values are indicated with black and white circles, respectively, and the vertical lines represent the interquartile range.Full size imageFig. 3: Global spatial variability in SST, SSS and SSD for coastal waters bordering mangrove forests under RCP 8.5.a, Global map showing the provinces (colour code and numbers) from the MEOW database28 used to investigate spatial patterns in mangrove coastal ocean water changes by 2100. b–d, Longitudinal gradient of the change in SST (b), SSS (c) and SSD (d) under RCP 8.5 in the AEP and the IWP mangrove bioregions; circles are coloured according to the MEOW province in which respective mangrove sites are located.Full size imagePredicted SSS changes exhibit an opposite trend in the AEP and IWP bioregions, with increased salinity in the AEP and reduced salinity in the IWP under global warming (RCP 2.6–RCP 8.5; Fig. 2b); this is reflected in contrasting SSD changes in both mangrove bioregions (Fig. 2c) and associated with predicted global changes in precipitation, with extensions of the rainy season over most of the monsoon domains, except for the American monsoon34. Under RCP 8.5, the spatially averaged change in mean SSS is +0.51 (±0.57) PSU in the AEP and −0.68 (±0.44) PSU in the IWP region. The maximum decrease in mean SSS (−2.01 PSU) is predicted for the Gulf of Guinea in the AEP bioregion (Fig. 3c and Supplementary Table 4). Within the IWP, the Western Indian Ocean region shows little or no changes in SSS, which contrasts with the pronounced freshening trends predicted in the eastern part of this ocean basin and the Tropical West Pacific (Figs. 1b and 3c). Increased freshening is predicted in the Bay of Bengal (SSS change: −1.17 ± 0.43 PSU), the Sunda Shelf (SSS change: −1.21 ± 0.29 PSU) and the Western Coral Triangle province (mean SSS change: −0.80 ± 0.17 PSU) (Fig. 3c and Supplementary Table 4). Within the AEP, salinity increases exceed +0.96 PSU in the Tropical Northwestern Atlantic, +0.80 in the Warm Temperate Northwest Atlantic and +0.68 in the West African Transition (Fig. 3c and Supplementary Table 4). The spatial heterogeneity in SSS across the global range of mangrove forests corresponds with observed changes in SSS35. Trends in SSD (Fig. 3d) strongly track changes in SSS (Fig. 3c) rather than SST. All RCP scenarios predict an overall decrease in SSD for both mangrove bioregions; however, the predicted decrease in SSD in the IWP region was a factor of 2 (RCP 6.0) and 2.5 (RCP 2.6, RCP 4.5 and RCP 8.5) stronger than in the AEP (Figs. 2 and 3d and Supplementary Tables 1–4).Propagule density values from our literature survey range from 1,080 kg m−3 for different mangrove species (Fig. 4 and Supplementary Table 5). The low densities reported for Heritiera littoralis propagules provide a strong contrast with the near-seawater propagule densities reported for Avicennia and members of the Rhizophoraceae (Bruguiera, Rhizophora and Ceriops). Floating characteristics of the latter may be particularly sensitive to changes in SSD. To illustrate the potential influence of changing ocean conditions on mangrove propagule dispersal, we considered threshold water density values (1,020 and 1,022 kg m−3) that are within the range where elongated propagules of important mangrove genera tend to change floating orientation (Fig. 4a). More specifically, we determined the ocean surface area with an SSD below or equal to these thresholds under different climate change scenarios (Fig. 5). Under RCP 8.5, the ocean surface covered by mangrove coastal waters (coastal waters bordering present mangrove forests) with a density ≤1,020 kg m−3 increases ~27% by 2100, notably more so in the IWP (~37%) than in the AEP (~6%) (Supplementary Table 6). A threshold of 1,022 kg m−3 results in increases of roughly +11% (global), +12% (IWP) and +8% (AEP) (Supplementary Table 7). Similar spatial patterns are observed for open-ocean waters within the global latitudinal range of mangroves (Fig. 5 and Supplementary Figs. 1 and 2).Fig. 4: Potential effect of future declines in SSD on mangrove propagule dispersal.a, Range of reported propagule density values for wide-ranging mangrove species and present and future range of SSD for coastal waters along the range of those mangrove species. Mangrove propagule data are extracted from the literature (Supplementary Table 5). H. lit, Heritiera littoralis; X. gra, Xylocarpus granatum; A. ger, Avicennia germinans; A. mar, Avicennia marina; B. gym, Bruguiera gymnorrhiza; C. tag, Ceriops tagal; R. man, Rhizophora mangle; R. muc, Rhizophora mucronata. Bottom part adapted from ref. 51. b, Conceptual figure of the potential effects of ocean warming and freshening on mangrove propagule dispersal. Ocean warming and freshening drive changes in SSD and may reduce the timeframe for opportunistic colonization. For a propagule with a specific density and floating profile under present surface ocean conditions, reduced SSD of coastal and open-ocean waters may reduce floatation time (shaded area) and hence, reduce the proportion of long-distance dispersers. For simplicity, the density of propagules is assumed to increase linearly over time, although the actual increase may be nonlinear.Full size imageFig. 5: Future changes in SSD.a–d, Spatial extent of coastal and open-ocean surface waters with a density ≤1,020 kg m−3 (a,b) and 1,022 kg m−3 (c,d), for present (2000–2014) (a,c) and future (2090–2100; RCP 8.5) (b,d) scenarios. Data are shown for surface ocean waters within the global latitudinal range of mangrove forests (between 32° N and 38° S). The two density thresholds considered are within the range of densities at which mangrove propagule buoyancy and floating orientation of several mangrove genera change, as reported in available literature. Black dots along the coast represent the global mangrove extent from the 2015 GMW dataset32. Magenta-coloured circles represent SSD values More

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Key delimitation trapping survey performance factorsTrap attractivenessThe performance of the current Medfly design was unexpectedly inferior to that of the leek moth even with a more vagile target insect, 2.8 times greater trap density in the core, and a grid size over three times larger. Despite all those factors, p(capture) for the leek moth grid with 1/λ = 20 m was 15 percentage points greater than that for Medfly at 30 days duration. Thus, trap attractiveness was the key determinant for delimiting survey performance, as it was for detection13.One straightforward way to improve p(capture) and the accuracy of boundary setting, while also cutting costs, would be to develop more attractive traps. Poorly attractive traps include food-based attractants48 and traps based solely on visual stimuli36. But developing better traps is difficult. Pheromone-based attractants generally perform best49, but these are unavailable for many insects. For instance, scientists have searched for decades for effective pheromones for Anastrepha suspensa (Loew) and A. ludens (Loew) without success50. Common issues include the complexity of components, costs of synthesis, and chemical stability.Trap densitiesAll else being equal, increasing the trap density will generally improve p(capture) for any survey grid, and intuitively this can help compensate for using less attractive traps. However, the impact of increasing density is limited when attractiveness is low13,47, and large surveys or grids with many traps can become prohibitively expensive51. The Medfly grid designers likely understood that the available trap and lure was not highly attractive, and used higher densities in inner bands to try to reach some desired (non-quantitative) survey performance level. By contrast, the designers of the leek moth grid used a (constant) density three times smaller, likely because the trap and lure were known to be relatively strong. Here, for both species, marginal ROI decreased as densities increased (Tables 2, 3). Hence, increasing densities has limited benefit, but may be useful when better lures are unavailable13.In that context, the use of variable densities in the Medfly grid is understandable. At its standard size, the survey grid would require 8,100 traps if the core trap density were constant (Table 1). The designers likely intuited that lower densities could be used in outer bands because captures there were less likely. However, doing so reduces the likelihood of detection in outer bands and could increase the possibility of undetected egress, especially with longer survey durations. As far as we know, natural egress has not been raised as a concern following the numerous Medfly quarantines that have used this survey grid over the years, in Southern California in particular52.Generally, however, we think the variable Medfly grid densities run counter to delimitation goals. Greater core and Band 2 densities have proportionally more impact on p(capture), but only a few detections in the core are necessary to confirm the presence of the population (Goal 1), and inner area detections probably contribute little to boundary setting (see below). Therefore, lower or intermediate densities (at most) may be optimal for the core when considering ROI. For the outer bands, increasing densities might improve boundary setting (Goal 2) and help mitigate potential egress, but the sizes of those bands already limit cost efficiency (Table 2), making greater densities less advisable. Our simulation results can help elucidate how to balance these interests to achieve delimitation goals while minimizing costs47.Grid size considerationsThe simulation results indicated that the standard survey sizes for these two pests were excessive. We have verified that empirically for Medfly using trapping detections data53. A 14.5-km grid has been widely used for many other insects in the CDFA (2013) guidelines10, such as Mexfly and OFF, and the same analysis indicated that those are also oversized for use in short-term delimitation surveys53. From the same analysis, the predicted survey radius for leek moth, with D = 500 m2 per day, would be 2,382 m, or a diameter of nearly 4.8 km, which matches the results here. Similarly, Dominiak and Fanson45 analyzed trapping data for Qfly and found that the recommended quarantine area distance of 15 km could be reduced to 3 to 4 km.Grids with radii larger than 4.8-km only seem necessary for highly vagile insects, those with D ≥ 50,000 m2 per day47. This should not be surprising. Small insect populations are unlikely to move very far31,54, especially if hosts are available20,39,55. The (proposed) short duration of a delimitation survey would also limit dispersal potential (see below). Many delimiting survey plans may be oversized, because they were developed before much dispersal research had been done37, thus uncertainty was high. Our dispersal distance analysis included species with a wide range of dispersal abilities, so it can be used generally to choose smaller survey grid radii53.Reducing grid sizes down to about 4.8-km diameters may have little impact on p(capture), since detections in bands outside that distance contributed little to overall performance. The cores of both the leek moth and Medfly grids accounted for 86 percent or more of overall p(capture). While core area detections will confirm the presence of the population, they are less useful for defining spatial extent. The furthest detections from the presumed source are usually used to delimit the incursion46,56 (although in our experience formal boundary setting exercises seem rare). Delimiting surveys may often yield few captures anyway, because adventive populations can be very small and subject to high mortality31. Because size reductions eliminate traps in proportionally larger outer areas, the impact on survey costs is substantial. Removing just the outermost bands of each grid would directly reduce costs by $11,200 for leek moth (400 traps) and by $7,488 for Medfly (288 traps; Table 1).Another reason for the large size of the standard Medfly grid may be that it was designed for monitoring and management in addition to delimitation57. Medfly quarantines end after at least three generations without a detection, so the surveys may last for months. The grid size was reportedly originally determined by multiplying the estimated dispersal distance by three (PPQ, personal communication), to account for uncertainty. This implies that the estimated distance was about 2,400 m per 30 days. Thus, the design may not have been built for the 30-d duration used here, but our recommended design is valid if a shorter delimitation activity without further monitoring is appropriate.Although it seemed too large for leek moth, an 8-km grid for delimitation could be appropriate for some other moths. For example, the delimiting survey plans for Spodoptera littoralis (Boisduval) and S. exempta Walker use this size9. S. littoralis is described as dispersing “many miles”, and S. exempta can travel hundreds of miles9, which clearly exceeds the described dispersal ability of leek moth. On the other hand, the survey plan for summer fruit tortrix moth (Adoxophyes orana Fischer von Röeslerstamm) also specifies an 8-km grid for delimitation but contains little information on dispersal, suggesting only that most movement is local8. Like leek moth, a 4.8-km grid for that species seems likely to be more appropriate.Limiting egress potential is probably the main consideration when setting survey size, but uncertainty about the source population location may also be a factor. Survey grids placed over the earliest insect detection may sometimes be off center from the location of the source population54. However, so far as we know for our agency, most adventive populations have been localized, based on post-discovery detections (PPQ, personal communication). Likewise, we have found53 and other researchers have found that dispersal distances for different species in outbreaks and mark-recapture studies are often less than 1 km58,59,60. That may often be the case for detection networks of traps (e.g., for high risk fruit flies), which increase the likelihood of capture before the population has had much time to grow and disperse. Here, we focused explicitly on localized populations, but allowed for uncertainty in the simulations by varying outbreak locations over one mile in the central part of the grid. If the outbreak population is very large and has extensively spread out (e.g., spotted lanternfly, Lycorma delicatula (White) in 201461), delimitation will not be localized, but “area-wide”2. The results here do not apply to area-wide outbreaks, and we are currently studying how to effectively delimit them.Optimizing delimitation surveysMany trapping survey designs in use were based not on “hard” science but on local experience62. Scientists have recognized the need for more cost-effective surveillance strategies63,64. Quantitatively assessing p(capture) in different designs for the same target pest allows us to determine grid sizes and densities that lower costs while maintaining performance. Results here demonstrated that the sizes and densities of these two survey grids could be optimized to save up to $20,244 per survey for the leek moth and $38,168 per survey for the Medfly. In practical terms, that means more than five leek moth surveys could be run for the cost of one standard design survey. Additionally, over seven Medfly delimitation surveys could be funded by the budget of one standard plan. The magnitudes of reduction seen here may be typical, since about 90 percent of the costs in trapping surveys are for transportation and maintenance related to traps65.Quantifying survey performance was not possible until very recently, so it has been little discussed in the literature5,66, and no standard thresholds exist. We think 0.5 may be a reasonable minimum threshold for the choice of p(capture), to try to ensure that population detection is “more likely than not”. Designs that aim to maximize p(capture) could be realistic with high attractiveness traps, but those designs seem very likely to have lower ROIs (e.g., Table 2). Even for the most serious insect pests, we think targeting near-perfect population detection during delimitation is likely not justified. Designs achieving p(capture) from 0.6 to 0.75 could be highly effective in terms of both costs and performance.Another potential area of improvement is grid shape. Circular grids perform as well as square grids but use fewer traps and less service area to achieve equivalent p(capture)47. Moreover, detections in the corners of a square grid are evidence that insects could have traveled beyond the square along the axes, resulting in uncertain boundary setting. Most published survey grids are square10,46, but many field managers tend to use approximately circular trapping grids in the field (PPQ, personal communication). The conversion to a circular grid with a radius of half the square side length reduces the area and number of traps by around 21 percent47. Our findings were consistent with that value.This new quantification ability also indicates that some delimiting survey designs in the U.S.A. may not be performing as well as expected47. For instance, the delimiting survey design for Mexfly uses approximately 31 traps per km2 in the core of a 14.5 km square grid11, but the traps are only weakly attractive (1/λ ≈ 5 m). In this scenario, p(capture) was only around 0.23 with a 30-d survey duration47. A much greater density ( > 80 traps per km2) could be used in the core to achieve p(capture) ≥ 0.5, but this may not be feasible depending on the survey budget.Technical and modeling considerationsExamining diffusion-based movement for these two insects in TrapGrid can give insight into why simulations indicated that smaller grids may be adequate47. The value of σ for Medfly after 30 days is only about 1,550 m. In a normal distribution, σ = 1,550 m gives a 95th percentile distance of 2,550 m, which is similar to the estimated distance above of 2,400 m. Over 90 days, σ = 2,700 m for Medfly, which gives a 95th percentile distance of 4,441 m, still much shorter than the grid radius of 7,250 m. A 95th percentile of 7,250 m requires σ ≈ 4,408 m, which equals t = 253 days. In addition, the maximum total distance (up to 39 days after detection) we observed in trapping detections data for Medfly in Florida was about 4,800 m53.The same calculations for leek moth give σ ≈ 490 m for 30 days, with a 95th percentile distance of only 806 m. That is half the length of the recommended shortened radius above of 2.4 km, and nearly five times shorter than the radius of the standard 8-km grid. A 95th percentile of 4,000 m requires σ = 2,432 m, which implies t = 740 days, which is about two years. Therefore, the leek moth grid is arguably even more oversized than the Medfly grid.The default capture probability calculation in the current version (Ver. 2019-12-11) of TrapGrid is not sensitive to population size32 and does not consider the effects of ambient factors (e.g., wind speed and direction, rainfall, temperature). Many other factors can also impact trapping survey outcomes, such as topography of the environment, availability of host plants, seasonality of pest, and population dynamics. These factors are not considered in the current version of TrapGrid. More

Study siteThe study was conducted at the Center for Applied Research on the Environment and Sustainability (CARES) at The American University in Cairo, New Cairo, Egypt (30°01′11.7″N 31°29′59.8″E) from 12/Nov/2019 until 31st/March/2020. The experiment was carried out in a greenhouse-controlled environment with temperatures ranging from 18 to 23 °C and relative humidity between 60 and 70% during the growing period.Experimental designThe proposed design starts by treating brackish water using RO membrane separation technology, powered by an on-grid 10 kW photovoltaic solar panel as shown in Fig. 1. The permeate (freshwater) from the RO facility is directed to the aquaculture units of capacity of 1 m3, where the fish effluents are used as irrigation water and as the sole source of fertilizers for the crops.Figure 1Schematic Integrated model design. T1 Deep water culture system without sand, T2 Sandponics system with sand from October, T3 Sandponics system with sand from Beni suef, T4 Sandponics system with sand from Fayoum.Full size imageThe study followed a completely randomized design with four variants, i.e., an aquaponic deep-water culture system (T1) and three sandponics systems (T2–T4). The three sandponics systems were established with different sand collected from different sand locations in Egypt during the period between September and October 2019.Initially, an exploratory field trip was set to six different locations in Egypt to collect sand samples for lab analysis aimed at sourcing the most suitable sand for the system under study with regards to both the physical and chemical parameters. These areas include Ismailia Governorate; 30°34′55.2″N 31°50′08.1″E, 6th October governorate; 29°54′49.8″N 31°05′51.5″E, Benu Suef governorate; 28°53′18.4″N 30°45′12.9″E, Al-Minya governorate; 28.725799, 30.630305, and two sites from Fayoum governorate; 29°05′07.4″N 30°49′39.9″E.From the six locations in Egypt, preliminary sand analysis was carried out, and sand samples were also collected for both physical and chemical lab analysis at the Soil and Water Lab at the Agricultural Research Center in Dokki, Egypt. Following a thorough technical, field, mechanical, and lab chemical evaluation of the six sand samples from six locations, three sand locations/types were selected for experimentation that seemed fit and suitable for the current study. The criteria parameters for the shortlisting of sand included water retention potential of the sand by the percolation process, testing the carbonates level in the soil, the turbidity of the sand, porosity percentage and drainage potential of the sand. The three locations included 6th October (T2), Benu Suef (T3), and Fayoum site 2 (T4). In the second week of November 2019, ten cubic meter tracks of sand from the three above locations were set to collect sand from these areas to the research facility at CARES where the experiment was carried out.The study was carried out with two systems/setups, i.e., an aquaponic Deep Water Culture (DWC) and SP systems. The DWC model comprises a 1 m3 fish tank, a settlement tank, a mechanical filter, a biological filter, three grow beds, and a drainage tank. This system being the most practiced aquaponics technique was considered as the control. Fish effluent water flowed from the fish tank to the settlement tank to filter big solid wastes through the mechanical filter to remove the smaller solid wastes and the biological filter for the nitrification process. Then filtered water continues to the grow beds, where overflow drains into the drainage tank and back to the fish tank in a closed system.On the other hand, the variable in the three IAVS systems is the sand source. This system comprises three independent set-ups: a 1 m3 fish tank, three grow beds, and a drainage tank. Fish effluents flowed from the fish tank directly to the sand grow beds where water was supplied through irrigation drip lines using diaghram emitters connected with valves to ensure uniformity of water application to each grow bed.All the fish tanks were installed with the same fish stock size of 30 Nile tilapia (Oreochromis niloticus) from an existing fish stock at the research center with an average initial weight of 244 g and the same amount of water, initially 850L per tank. The fish was sourced from an already existing aquaponics system at the research center to avoid any transportation stress effects and related shocks on the small fish, leading to a lot of mortality cases. The fish were fed 3–4 times daily with commercial pellets containing 30% proteins, 5% crude lipid, 6% crude fiber, 13% Ash, and 9% moisture content supplied by Skretting Egypt. The feeding pattern and frequency were according to the fish body biomass percentage of 2–3% depending on the growth stage and upon reaching satiation.DesalinationThe experiment was entirely run with desalinated water produced from a desalination facility at the center. The desalination technology used was Reverse Osmosis (RO); in batch mode; using a Sea Water Pump with Energy Recovery Unit (model Danfoss-APP1.0/APM1.2). The RO membrane used is Hydraunatic SWC5-4040, from Lenntech company with an average salt rejection of 99.7%. Three modules were connected in a series arrangement (3 Pressure Vessels each equipped with a single module). Synthesized brackish water was prepared by dissolving industrial grade sodium chloride (sea salt) from El-Arish Governorate, Egypt. The salt chemical properties are presented in Table 1. Feedwater salinity was 10 mg/L, with an equivalent osmotic pressure equal to 8.61 bars. The osmotic pressure was calculated using Van’t Hoff relation. Permeate Total Dissolved Solids (TDS) was 192 mg/L, and brine TDS was 13.1 g/L as shown in Table 2.Table 1 Chemical properties of the used salt.Full size tableTable 2 Chemical properties of water samples used.Full size tableThe average pure water flux is 9.5 LMH and was calculated by dividing the permeate volume by the product of membrane surface area and time. Each batch run produced around 4 m3 of permeate, which was enough to irrigate the designated plant beds. The estimated average permeate recovery for the RO process is 22% and salt rejection exceeded 98.7%. The differential pressure between membrane inlet and outlet was equal to 1 bar, where membrane inlet pressure was 16 bars, and the outlet was 15 bars. The RO process operated at an average transmembrane pressure equal to 16 bars and an average permeate and brine flow rates equivalent to 3.49 and 12.41 Lpm, respectively. All experiment runs were performed at 25 °C.Plant materials and cultivation practiceSwiss chard bright lights (Beta vulgaris subsp. cicia) seeds were imported from Seed kingdom seed company in the USA. Seeds were sown in ¼ inch holes in a seed starting mix containing perlite and vermiculite and irrigated with a hand mist sprayer daily to keep the growing media always moist. Sowing was done on the 12th of November 2019, and seedlings were transplanted when they were 40 days old. Seedlings were transplanted into raised grow beds made of fiberglass material measuring 1.8 × 1.2 × 0.6 m for each of the four systems. The beds were raised off the ground by 0.5 m to allow drainage water from the bed to be collected and circulated back to the fish tank. Each bed was constructed with a drainage pipe at the bottom covered with a mesh net to prevent water blockage by the sand. Also, a 5 cm layer of small gravel was uniformly laid at the bottom of the beds to facilitate drainage, followed by sand with a height of 50 cm.In the IAVS systems, plants were irrigated using manually punched diaphragm emitters, and the irrigation flow rate was controlled using small plastic valves at the start of every irrigation tube. Emitters were installed in drip tubing at a 30 cm distance as well the tubing lines were also placed 30 cm between each other. Seedlings were transplanted 5 cm away from the emitters at 30 cm between rows and 30 cm within the row. Since the water was pumped with submersible pumps to the grow beds, regulatory pressure valves were installed in between the pump and the main irrigation line, and then water flows through the emitters into the row furrows. Water would then saturate in the sand and eventually drain at the bottom into drainage tanks and pumped back to the fish tanks.To maintain the water quality, two full cycles of water recirculation were run every day. Every irrigation cycle recirculated 25% of the fish tank, and complete drainage was allowed for a maximum of two hours. Plants were harvested upon reaching maturity for three cuts, except with the T1, which could not grow back after the second cut. Plants took 52 days from transplanting to reach the first cut, 20 days from cut 1 to cut 2, and as well 23 days from cut 2 to reach cut 3. Measurable crop parameters included plant height at harvesting/cutting, leaf area, number of leaves per plant, chlorophyll content, fresh weight per plant, and nutrient composition. Since the focus of SP is on the crops, fish were only measured to monitor their relative growth in terms of weight gained at harvesting/cutting time.Measurement of crop parametersPlants were cut 5 cm above the soil surface, and agronomical trait measurements from a representative sample of 12 plants per replicate were taken as follows.Plant heights were taken using a foot ruler and averages determined. Leaf number was obtained as the number of leaves counted per plant and averages determined. Leaf area was calculated according to the equation reported by Yeshitila and Taye16.$${text{Leaf}} , {text{ Area }}left( {{text{cm}}^{{2}} } right) = , – {422}.{973} + { 22}.{752}0{text{L }}left( {{text{cm}}} right) , + { 8}.{text{31W }}left( {{text{cm}}} right)$$where L and W represent the leaf length and Leaf width respectively, − 422.973 is a constant relating to the shape of the leaf of Swiss chard developed by the author under citation.Chlorophyll content was measured using MC-100 chlorophyll meter from Apogee Instruments, Inc, and data was expressed as SPAD averages. Fresh weight was measured using a digital weighing balance and data expressed as g/plant.Sand testSand samples were obtained and sent for analysis at the Soils, Water and Environment Research Institute, Agricultural Research Center, Giza, Egypt. The Electrical conductivity (EC) values were measured from the sand paste extract; pH values were taken from sand suspensions at ratio of 1:2.5 as described by Estefan17. The available nitrogen in the sand sample was extracted using potassium chloride (KCl) as an extractable solution with the ratio of (5gm sand to 50 ml KCl) and determined using the micro- kjeldahl method. Available potassium was determined using a flame photometer, and the other elements in the sand sample were determined by using inductively coupled plasma (ICP) Spectrometry (model Ultima 2 JY Plasma)18,19. The physical and chemical characteristics of the used sand are presented in Table 3.Table 3 (a): Chemical analysis of field sand samples, (b): Available macro, micronutrients, and heavy metals content of the sand samples.Full size tableWater analysisEvery 15 days, a measured amount of desalinated water was added to a standard mark of 850L in the fish tanks to compensate for the consumed amount of water in the system. Fish water quality parameters such as water temperature, pH, and dissolved oxygen (DO) was closely monitored using automated digital Nilebot technologies by Conative labs to fit the ideal required levels as reported by Somerville et al.20. In contrast, ammonia, nitrite, and nitrate were adjusted using an API test kit every week. These parameters’ recorded values were as follows: water temperature ranged between 25 and 28 °C, DO range between 6–7 mg/L, and pH between 6.5 and 7.0. Ammonia levels were kept below 1 mg/L. Elements in water samples were determined according to EPA methods18 using inductively coupled plasma (ICP) Spectrometry (model Ultima 2 JY Plasma) as presented in Table 4.Table 4 Water sample analysis for the different systems’ fish tanks and sump tanks.Full size tableNutritive composition analysisAccording to Official methods of analysis from the association of official analytical chemists (A.O.A.C) (1990), moisture content and Vitamin C were determined. Vitamin A was determined according to the procedures described by Aremu and Nweze21. Briefly, 100 g of the sample were homogenized, from which 1 g was obtained and soaked in 5 mL methanol for two hours at room temperature in the dark for complete extraction of a pro-vitamin A carotenoid, β-carotene. Separation of the β-carotene layer was achieved through the addition of hexane to the sample, and moisture was removed using sodium sulphonate. The absorbance of the layer was measured at 436 nm using hexane as a blank. β-carotene was calculated using the formula:$$beta {text{-carotene }}left( {{mu g}/{1}00{text{ g}}} right) , = {text{ Absorbance }}left( {text{436 nm}} right) , times {text{ V }} times {text{ D }} times { 1}00 , times { 1}00/{text{W }} times {text{ Y}}$$where: V = total volume of the extract; D = Dilution factor; W = Sample weight; Y = Percentage dry matter content of the sample.Vitamin A was then determined according to the concept of Retinol Equivalent (RE) of the β-carotene content of the vegetables using the standard conversion formula. Total hydrolyzable carbohydrates were determined as glucose using phenol–sulfuric acid reagent as described by Michel22.Vitamin C content was determined using dichlorophenol indophenol reagent. As such, 10 g of fresh leaf tissues, were crushed using a motor and pestle in the presence of 10 ml metaphosphoric acid 6% (Merck). This was followed by centrifugation at 4000×g for 5 min at 4 °C. Five mL of the supernatant were transferred into an Erlenmeyer flask, and 20 mL of 3% metaphosphoric acid were added. The extract was titrated by dichlorophenol indophenol (Sigma-Aldrich) until a rose color was observed. Vitamin C (mg/100 g FW) was then calculated and based on the standard curve of l-Ascorbic acid (Merck) concentrations.For the determination of protein and mineral content, 0.5 g of dried samples were digested using sulfuric acid (H2SO4) and hydrogen peroxide (H2O2) as described by Cottenie23. From the extracted sample, the following minerals were determined:Nitrogen was determined according to the procedures described by Plummer24. Briefly, 5 mL of the digestive solution was distilled with 10 mL of sodium hydroxide (NaOH) for 10 min to obtain ammonia. Back titration was then used to determine the amount of nitrogen present in ammonia. Protein content was calculated by multiplying total nitrogen by 6.25 according to methods of AOAC25.Phosphorus content was determined calorimetrically (660 nm) according to the procedures described by Jackson26. Potassium, Calcium, and Sodium were determined against a standard using a flame-photometer (JEN way flame photometer) as described by Piper27. Magnesium (Mg), Copper (Cu), Manganese (Mn), Zinc (Zn), and Iron (Fe) content were determined using Atomic Absorption Spectrophotometer, Pyeunican SP1900, according to methods described by Liu28.The moisture percentage of leaf samples was determined by weighing the fresh weight for each sample (Fw), then dried for 72 h at 80 °C. The dry matter weight was record as Dw. The leaf water content was then calculated as the following:$${text{Moisture}};{text{ content }}left( % right) , = , left( {{text{Fw}} – {text{Dw}}} right) , /{text{ Fw}} * {1}00$$Statistical analysisStatistical comparisons among means of more than two groups were performed with analysis of variance (ANOVA) using SPSS V22, and the difference in means was analyzed by Tukey’s test at α = 0.05. Statistical differences were considered significant at P ≤ 0.05 in triplicates and data expressed as mean ± S.D.Plant materialAll plant materials and related procedures in this study were done in accordance with the guidelines of the Institutional Review Board of the American University in Cairo and the Ministry of Agriculture and Land Reclamation in Egypt.Ethics approvalThis study followed the guidelines and approval of Committee of Animal Welfare and Research Ethics, Faculty of Agriculture, Kafrelsheikh University, Egypt. More

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