Introduction

Every year, French agriculture and agribusiness produce more than 300 million tons of organic effluents, of which 26 million tons of slurry and 0.8 million tons of manure spread per year in France come from pig farming (Loyon, 2017). These effluents represent considerable sources of nutrients and energy. However, they can also represent sources of air, water and soil pollution at both local and global levels, with harmful consequences on the environment and on animal and human health (OneHealth concept). The challenge is therefore to make better use of the resources present in effluents (nutrients, energy) in order to reduce their environmental impact and to use their agronomic, energetic and economic potential within the framework of a circular bioeconomy.

Because of their low absorption rate by pigs and their beneficial effects on health and growth performance, zinc (Zn) and copper (Cu) are fed in quantities higher than required. Pig's effluents can then contain quantities of Zn and Cu between 4 and 10 times higher than the needs of the plants when agronomic inputs are reasoned on the basis of nitrogen (N) or phosphorus (P) contents (Jondreville et al., 2002; Revy, 2003). As direct application is the main way of using effluents, these elements are therefore applied in excess and accumulate in soils (Coppenet et al., 1993; L'Herroux et al., 1997; Jensen et al., 2018), leading to risks of toxicity for soil microorganisms (McGrath et al., 1995) and for plants (McGrath, 1981; Jondreville et al., 2002; Revy, 2003). Furthermore, the use of pharmacological doses (see Box 1) of Zn in piglets would contribute to the development of bacteria resistant to antibiotics (Jensen et al., 2018), which is also an important issue for their reduction.

In order to limit these various risks, both for the environment and for animal and human health, European regulations are providing an increasingly strict framework for the quantities of Zn and Cu that can be added to animal feed (EU-2016/1095; EU-2018/1039), with nutrition being the main lever for reducing the flows. There are also regulations on the amount spread of these minerals, and specifications or standards on the Zn and Cu content of organic fertilisers and amendments. Furthermore, the various effluent treatment technologies that are being developed (composting, aerobic and anaerobic digestion, etc.) can lead to an increase in the concentration of these elements, particularly in relation to dry matter (DM) (Hsu and Lo, 2001; Legros et al., 2017). The knowledge and management of these flows in a context of diversification of effluent recovery procedures are thus important issues.

In this review, Zn and Cu flows and their fate are characterised along the continuum of feed-animal-manure-treatment-soil. The first part refers the use of Zn and Cu in pig feed and the needs of pigs. The release of these elements in effluents and their behaviour in the different pig effluent management systems are described in the second part. Finally, the impact of the return to the soil of products from pig farming and their Zn and Cu composition on the environment are discussed. Due to the limited data available on sows, this review focuses on piglets and fattening pigs.

1. Zinc and copper in pig feed

Zn and Cu are considered as essential minerals in pig nutrition. Their bioavailability in the raw materials of the basic feed ration is generally not sufficient to cover the physiological needs (Männer, 2008). Therefore, they are supplemented in the feed in quantities ranging from nutritional levels, in order to cover the animals' needs, to supra-nutritional levels (see Box 1) to improve growth or digestive health. However, as their retention is very low, the majority of them are found in the faeces. Reducing the dietary intake of Zn and Cu is therefore the main lever for reducing their quantity in effluents and their potential impact on the environment, while ensuring that they do not affect the animal's growth or health. To do this, it is essential to better understand their function and fate in the body, which influence their bioavailability and therefore their absorption and excretion.

1.2. Pig requirement

The requirement for an element is the amount needed to enable its body to perform all its physiological functions (Revy, 2003; Schlegel, 2010). It can vary according to the response criteria used to define it. As the satisfaction of the physiological requirement of animals depends on the bioavailability of minerals in the feed (National Research Council, 2012), the definition of the requirement must also consider the forms of intake. In practice, it is necessary to know the Zn and Cu requirements and their evolution with the age of the animal, in order to adapt the feed as best as possible and thus reduce the risks of deficits, which are harmful to performance, or excesses, which are harmful to the environment.

a. Assessment of piglet and growing pig requirement

Two main approaches are used to assess mineral requirement, one empirical and one factorial (Schlegel, 2010), the former being the most common for trace metals (TM). This consists of experimentally testing the effects of a gradual increase in the amounts of Zn and Cu in the feed in order to evaluate the response of several zootechnical or physiological parameters. Statistical treatment of the results by a linear plateau model is then generally used to describe the response and define the breakpoint requirement (Kirchgessner, 1993).

A specific and representative response of one of the animal's functions for Zn or Cu must be observed in order to determine a criterion as characteristic of the requirement (Schlegel, 2010). For Zn, the most representative criterion generally used is its concentration in bone or plasma; for Cu, this criterion is its concentration in the liver (Jongbloed, 2010).

This is illustrated in figure 1 for Zn in post-weaning piglets, based on different literature references. Depending on the level of intake, we can define i) a deficiency zone (in red) in which the intake is not sufficient to maintain an adequate plasma level; ii) a homeostasis zone (in green) in which the plasma level remains constant thanks to the implementation of different homeostasis mechanisms (storage, excretion...) and iii) a zone of excess (in blue) in which the plasma concentration remains constant.) and iii) a zone of excess (blue) in which the plasma concentration increases with food intake, the homeostasis mechanisms being probably saturated. The breakpoint between the red and green zones defines the nutritional requirement, whereas the breakpoint between the green and blue zones indicates the appearance of a risk of excess associated with a drop-in intake.

Figure 1: Evolution of plasma Zn concentration in post-weaning piglets as a function of the Zn content of the feed (Hahn and Baker, 1993; Hill et al., 2001; Revy, 2003).

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The National Research Council (2012) gives different references for Zn requirements for pigs depending on the growth or physiological stage of the animal. For weaners the requirement is estimated at 80 mg Zn per kg feed. For fattening pigs, the average requirement estimated in a basic ration by the NRC (2012) is about 50 mg.kg^-1 DM of feed. This requirement may vary according to different criteria that modify the bioavailability of Zn: the nature of the feed, its calcium (Ca) and phytate content, and the incorporation of phytase (Revy, 2003; Spears and Hansen, 2008).

The strict requirement for piglets for Cu is estimated at 5-6 mg.kg^-1 DM of feed and does not exceed this value for later physiological stages (National Research Council, 2012). However, this requirement does not consider the beneficial effects of higher supplementation levels on growth performance or digestive health at certain critical phases of the animal's life, such as weaning, or for breeding sows As for Zn, the Cu requirement may vary according to different criteria that modify the bioavailability of Cu, such as the addition of more Zn in the feed.

b. Deficiency effect

The main symptoms of Zn deficiency are initially loss of appetite, the onset of diarrhoea and growth retardation. Anorexia is one of the first signs observed and leads to reduced femur size and strength in deficient piglets (Suttle, 2010). Emergent diarrhoea is due to reduced enterocyte turnover (Revy, 2003). In addition, a decrease in plasma or serum Zn level is observed as well as alkaline phosphatase and albumin (Suttle, 2010; National Research Council, 2012). However, animals respond to Zn deficiency by regulating the various intestinal Zn transporters, but this is not sufficient to cover the premature decline in plasma and serum Zn concentration in deficient piglets (Suttle, 2010). The most prominent symptom of Zn deficiency and the last sign to appear is a hyperkeratinisation of the skin called parakeratosis (Suttle, 2010; National Research Council, 2012).

Cu deficiency in pigs are rarely observed and can occur in weanlings fed about 100 mg.kg^-1 MS Fe, 130 mg.kg^-1 MS Zn and 2 mg.kg^-1 MS Cu (Jondreville et al., 2002). Apparent signs of Cu deficiency are anaemia, limb bowing, the occurrence of spontaneous fractures and heart and vascular problems and depigmentation (National Research Council, 2012). Recent results (Dalto et al., 2021), however, indicate a risk of Cu deficiency developing in the case of Zn intake at pharmacological doses (2500-3000 mg.kg^-1 DM), confirming the reduction in plasma Cu observed by Hill et al. (2001) in this situation (see section 1.3.d).

c. Excess effect

Zn is not very toxic for most mammals. However, an excess of Zn (see Figure 1) in the feed leads to a decrease of its absorption, an increase of its storage in the bones and in the enterocytes after binding to metallothionein and therefore an increase in tissue turnover and endogenous Zn secretions. At pharmacological dosage (see Box 1), excess Zn intake can be accompanied by a reduction in feed intake and therefore in growth rate (Hahn and Baker, 1983). An alteration of Cu and Fe metabolism can also happen, with risks of deficiencies if the excess situation is prolonged (Dalto et al., 2021). Moreover, the susceptibility of the animal to excessive Zn also depends on the Ca, Fe, Cu and cadmium content of the feed (Revy, 2003).

Situations of Cu poisoning are rare in pigs (Jondreville et al., 2002). However, a long-term distribution of more than 250 mg.kg^-1 DM of Cu can lead to a toxic effect and the symptoms observed are a reduction in haemoglobin levels and jaundice (National Research Council, 2012). This can lead to a reduction in Fe storage in the liver and therefore anaemia, due to a negative reaction of Cu on Fe, which has limited absorption (Jondreville et al., 2002). The tolerance of pigs to high dietary doses of Cu depends on the supply of Zn in the feed and the interactions of Zn and Cu with metallothionein (Jondreville et al., 2002).

Finally, the capacity of a feed to fulfill the Zn and Cu requirements of pigs and the potential toxic effect of these elements depend on their concentration but also on their bioavailability (Hahn and Baker, 1983; Suttle, 2010); the absorption rate and the corresponding efficiency modify the total TM requirements of the animals (Männer, 2008).

1.3. Dietary intake of zinc and copper

a. Regulation

The European regulations concerning the maximum permitted levels of Zn and Cu in pig feed have changed since 2003 (Figure 2).

Figure 2: European regulations on maximum allowed levels of zinc and copper in feed for growing pigs (Regulation (EC) No 1334/2003; Regulation (EU) 2016/1095; Regulation (EU) 2018/1039 of 23 July 2018; Regulation (EU) 2019/4; Regulation (EU) 2019/6).

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From 28 January 2022, the use of zinc oxide (ZnO) at pharmacological level was banned in Europe. This regulation aims to reduce the excretion of Zn in faeces (Revy, 2003) and its accumulation in the soil, and to limit the development of antibiotic resistance in animals and humans to which high intakes of Zn contribute (Ciesinski et al., 2018).

b. Composition of raw materials

The basal diat of the pigs is mainly composed of cereals (mainly maize and wheat, but also barley, triticale, etc.), oil cakes (mainly soya, but also rapeseed and sunflower), oilseeds (peas, beans, etc.), cereal co-products (bran, middlings, draff, etc.), and vitamin mineral supplements (feed additives).

Cereals and protein crops contain on average 20-30 mg.kg^-1 DM of Zn, cereal co-products have a higher Zn content (between 70 and 90 mg.kg^-1 DM, Figure 3). Oilseeds and oilcakes contain between 30 and 90 mg.kg^-1 DM of Zn (figure 3).

Cereals, their co-products, oilseeds and oilcakes contain between 5 and 15 mg of Cu per kg DM (Figure 3).

Figure 3: Zn content of the raw materials of the basic pig ration as a function of their Cu content (after INRA-AFZ, 2004)

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c. Different forms of supplementation

Zn supplementation is essential in pig diets in order to cover the animal's needs, as the intake from raw materials alone is not sufficient. Moreover, the variation in the bioavailability of dietary Zn due to the different components of the ration must also be considered (Revy, 2003).

Contrary to Zn, a non-supplemented feed can theoretically cover the Cu requirement of growing pigs (about 6 mg.kg^-1). However, in practice, Cu supplementation in feed is usually applied to consider the imprecision of the total requirement estimation and to counterbalance the antagonistic effects of some elements in the ration, which affect the bioavailability of Cu and thus the requirement of the pig. Cu supplementation in the feed is used as a safety margin (Jondreville et al., 2002).

Zn and Cu, when added as additives, can come from different sources. Sulphates (ZnSO₄ and CuSO₄) are generally considered as the reference sources in comparative bioavailability studies. However, the fact that these sources are highly soluble means that they can also be more easily bound to other elements in the ration, particularly phytates in many feedstuffs, thus reducing their bioavailability (Revy, 2003). There are other sources of supplementation (oxides, chlorides, chelates) which have very variable physicochemical characteristics and bioavailability. In practice, ZnO and CuSO₄ are the most frequently used sources for animal feed (Revy, 2003).

d. Bioavailability of copper and zinc from feed and factors of variability

Two experimental approaches exist to measure the bioavailability of a mineral. The first is to measure the response of different physiological criteria to increasing intakes of the mineral, while remaining below the requirement (red area in Figure 1). The second is to measure this response with intakes well above requirement and measure their accumulation in the blood or in different tissues such as bone or liver (Spears and Hansen, 2008). In both cases, these responses are assessed relative to a reference source (CuSO₄ and ZnSO₄) whose bioavailability is set at 100%. Bioavailability is then calculated by comparing the response slopes of the different sources tested. Generally speaking, the bioavailability of minerals is influenced by the type of mineral supplementation provided, by factors such as the physiological status of the animal or its mineral status, and also by interaction effects with other agonistic or antagonistic elements of the ration (O′Dell, 1989; Männer, 2008). Different types of possible interactions are the formation of non-absorbable complexes in the gut, competition between cations for the transport of non-specific divalent cations or even the induction of proteins by non-specific metals (Suttle, 2010). The difference in bioavailability of Zn and Cu between sources could be largely explained by these phenomena that affect absorption, as well as their different physicochemical properties that affect their solubility.

Phytates form non-absorbable complexes with Zn, which reduces the bioavailability of Zn (Suttle, 2010). The hydrolysis of phytates allows the release of Zn, but monogastric animals do not naturally produce phytase, or in very limited quantities. The addition of microbial phytase to the ration can therefore improve Zn availability. Revy et al (2004) showed that 700 units of phytase can replace 32-43 mg of Zn in the form of sulphates. Similarly, Bikker et al. (2012) estimated that 500 phytase units can replace 27 mg of Zn as sulphate. According to the meta-analysis conducted by Schlegel and Jondreville (2011), only the Zn naturally present in the feed is influenced by phytates and phytase, and not the Zn added to the ration in the form of mineral or organic supplements. This antagonistic effect of phytates is increased after adding Ca to the ration (Suttle, 2010). A phytate-Ca-Zn complex is formed and precipitates Zn in the digestive tract (Revy, 2003). The antagonistic effect of phytates on Zn is all the more important in young animals, which are fed diets providing more Ca than required (Suttle, 2010).

Cu is less affected by phytates and Ca of the ration, due to its higher affinity for free amino acids, with which it forms chelates, allowing Cu to retain its solubility (Jondreville et al., 2002). However, soluble sources of Cu can interact in the digestive tract with phytates and form zinc-calcium-copper-phytate or copper-calcium-phytate complexes (Oberleas, 1973), which are resistant to phytase hydrolytic activity.

Zn has an antagonistic effect on Cu, i.e. it inhibits its absorption. This effect is due to the inducing effect of Zn on the synthesis of metallothionein, which have a strong affinity with Cu, preventing its transfer to the intestinal epithelium (Revy, 2003).

2. Zinc and copper excretion by pigs and manure management

Zn and Cu are few retained by animals. They are therefore mainly excreted in excreta, mainly in faeces. The concentrations of these TM can then constitute factors limiting the agronomic use of these excreta. Controlling these levels is therefore important to optimise input strategies according to the effluent utilisation pathway.

2.2. Feed strategy and reduction of copper and zinc excretion

There are different dietary levers to reduce Zn and Cu excretion. It is indeed possible to vary either the form of supplementation, or the amount of Zn and Cu added to the feed, or both simultaneously. Using the approach described by Dourmad et al (2013), we calculated the Zn and Cu excretion of pigs between weaning and slaughter (from 8 to 118 kg PV) under different feeding assumptions regarding Zn and Cu levels and considering the evolution of the EU regulations since 2003 and the perspectives for the future. The results presented in figure 4A and 4B clearly show that the regulatory changes have led to a strong reduction in excretion for both Zn and Cu.

Figure 4: Influence of changes in regulations on zinc (Zn) and copper (Cu) content in feed on excretion and effluent content: amount of Zn (A) and Cu (B) excreted per pig between 8 and 118 kg live weight (LW), and average Zn (C) and Cu (D) content of manure and digestate from anaerobic digestion; GP, 0GP: with or without Zn as a growth promoter

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Perspectives are 100 ppm Zn for post-weaning piglets and 80 ppm Zn for the fattening pig, and 100 ppm Cu for the first post-weaning period, 80 ppm Cu for the end of post-weaning and 15 ppm Cu in the feed for fattening pigs.

For Zn and with the new regulation from 2022, a 63% reduction in excretion per pig between 8 and 118 kg of PV is observed compared to the situation before 2003 (31.5 vs 85.4 g/pig). For Cu the evolution is even more severe with the current regulation the excretion is reduced by 80% compared to the situation before 2003 (7.6 vs 38.1 g/pig between 8 and 118 kg PV).

This evolution of excretion was also accompanied by a strong reduction in the content of the excreta. For Zn, this content has fallen from 2,100 to 1,000 mg.kg^-1 DM and for Cu from 1,100 to 250 mg.kg^-1 DM. Further reduction prospects can be planned (Figure 4.C and 4.D), but they require further knowledge of animal needs and bioavailability of sources (Dourmad et al., 2013).

Figure 5: Zn and Cu content of manure (mg.kg-1 DM) as a function of the Zn or Cu content of the feed and the sources of input (after Levasseur and Texier, 2001; Creech et al., 2004; Van Heugten et al., 2004; Hernández et al., 2008; Liu et al., 2016).

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Dark blue: pregnant sows; light blue: lactating sows; green: post-weaning piglets; yellow: fattening pigs.

Diamond: no supplementation; Round: inorganic source of Zn or Cu; Triangle: organic source of Zn or Cu.

The composition of manure also depends on the physiological stages of the animals (figure 5). The Zn and Cu contents relative to DM, depend both on their content in the feed and on the composition of the feed, in particular its fibre content which influences OM digestibility. Many studies have investigated the effect of modifying the Zn and/or Cu content of the feed on their concentrations in the manure. After reduction of the levels of these TM in the feed (to about 70 mg.kg^-1 DM of Zn and 10 mg.kg^-1 DM of Cu, compared to levels complying with European regulations, i.e. 150 and 25 mg.kg^-1 DM respectively), no effect on the performance of fattening pigs or on carcass quality was observed (Paboeuf et al, 2001; Creech et al., 2004; Van Heugten et al., 2004; Liu et al., 2016; Villagómez-Estrada et al., 2020b). All these studies show that the reduced Zn and Cu feed are effective in reducing Zn and Cu concentrations in manure. Different mineral sources have also been tested to decrease Zn and Cu excretion (Figure 5).

2.3 Pig manure management and impact on zinc and copper

Manure management is an opportunity to better control the distribution of TM in pig manure before it goes back to the soil and thus reduce its environmental impact. Effluent treatment does not eliminate metals but influences their concentration, due to OM degradation and/or phase separations, and their speciation (see Box 1), due to different chemical and biochemical mechanisms (adsorption, chemisorption, precipitation...) (Couturier, 2002).

a. Pig manure management systems

Although the most common way of managing manure is storage followed by spreading (Nicholson and Chambers, 2008), almost 12% of French pig farms were already using an alternative or complementary manure treatment technique to 'storage - spreading' in 2008 (Loyon, 2017). The treatments applied can be grouped into different categories: mechanical (phase separation), biological (aerobic treatment or anaerobic digestion, composting), chemical and thermal. These different technologies can be combined in different ways and are part of the overall manure management system (Figure 6 and Table 2).

Figure 6. Livestock manure and its possible fate (after Møller et al., 2007 Nicholson and Chambers, 2008; Loyon, 2017).

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Table 2. Main manure management pathways for pigs (after Møller et al., 2007; Nicholson and Chambers, 2008; Loyon, 2017) .

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Type of treatment	Type of effluent treated	Products obtained	Next possible treatment
Mechanical Treatments
Phase separation Sedimentation, screw compactor, centrifugation	Slurry Liquid phase from a separation in a building Digestate from methanisation Purified effluent	Solid phase	Anaerobic digestion Composting Thermal treatment
		Liquid phase	Aerobic treatment
Biological treatments
Aerobic treatment	Slurry Liquid phase from a separation in a building Liquid phase from phase separation	Purified effluent	Phase separation
Anaerobic digestion Mesophilic (35-40°C), hermophilic (55°C)	Slurry Solid phase from a building separation Solid phase from phase separation Manure	Digestate	Phase separation Heat treatment
Composting	Solid phase from building separation Solid phase from phase separation Manure	Compost
Thermal treatment Incineration, gasification thermal gasification, drying drying, evapo-concentration	Solid phase from a building separation Solid phase from phase separation Manure	Bottom ash Fly ash
Chemical treatments	Slurry

b. Effect of treatments on zinc and copper in effluents

Table 3 summarises the main effects observed on Zn and Cu during the treatments.

Table 3. Effects of different pig manure management and treatment on Zn and Cu elements.

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Management/Processing	Treatment effects on Zn and Cu	Parameter to be taken into account	References
Storage	- Increase in the influence of the distribution of Zn and Cu between the different classes of particles: the binding of metals to the particles	Type of storage Duration Temperature	Popovic and Jensen (2012)
Phase separation	- Concentration of Zn and Cu in the solid phase as a function of separation efficiency	Scraping method in the building	Loussouarn et al (2014) Likiliki et al (2020)
		Type of separator	Møller et al (2007) Nicholson and Chambers (2008) Pantelopoulos and Aronsson (2020)
Aerobic treatment	- Increased concentration of Zn and Cu, relative to OM and DM - Redistribution of Zn and Cu between solid (screenings) and liquid phases (supernatant and sludge)	Type of separators	Levasseur (2003) Béline et al (2004)
Methanisation / Anaerobic digestion	- Increase in Zn and Cu concentration, relative to OM and DM - Increase in the share of Zn and Cu in solid particles - Increased concentration of Zn and Cu of Zn and Cu in the poorly bioavailable fraction available fraction(unrecognized symbol) Decrease in the phytoavailability of these elements	Temperature pH Retention time hydraulic Oxidation-reduction reduction	Marcato (2007) Marcato et al (2009) Amaral et al (2014) Matheri et al (2016) Legros et al (2017) Yang et al (2020)
Thermal treatment (incineration)	- Cu accumulation in bottom ash - Concentration of Zn and Cu in particles > 30 µm		Møller et al (2007) Kuligowski et al (2008)
Composting	- Increased concentration of of Zn and Cu, relative to OM and DM - Increase of soluble part at the beginning of the process and then - Decrease of Cu in this form at the end of the composting process - Decrease in bioavailability factor of Zn and Cu	Moisture pH Nitrogen content Dissolved organic carbon organic carbon content Humic matter content	Hsu and Lo (2001) He et al (2009) Santos et al (2018) Li et al (2019)

- Phase separation in buildings

Phase separation directly in the building, e.g. by the V-scraping process, is fairly recent and the characterisation of the products obtained is still few reviewed. Zn and Cu follow mostly the solid fraction (between 90 and 95%) and this fraction is mainly treated by composting or anaerobic digestion. (Loussouarn et al., 2014; Likiliki et al., 2020).

- Duration and type of storage

Few studies exist on the effect of storage on Zn and Cu. According to Popovic and Jensen (2012), storage has no significant effect on Zn and Cu concentrations but increases the binding of Zn and Cu to less soluble solid particles.

- Mechanical phase separation

As only about 10% of Zn and Cu are soluble, these elements are mainly found in the solid fraction after separation from the slurry (Nicholson and Chambers, 2008). This redistribution will however depend on the type of separator or combination of separation processes used (Møller et al., 2007; Pantelopoulos and Aronsson, 2020). The separation of Zn and Cu to the solid phase is highest for chemical treatment with filtration (99.5% Zn and 94.9% Cu in the solid phase) followed by centrifugation (38.1% Zn and 31.1% Cu) and then screw compacting (7.1% Zn and 6.6% Cu) (Møller et al., 2007; Popovic et al., 2012; Pantelopoulos and Aronsson, 2020). Several studies evaluate the effect of combining different types of separation with an aerobic biological treatment plant (Levasseur, 2003; Beline et al., 2004). The separation results in lower concentrations in the liquid phase, thus reducing the annual soil load of Zn and Cu after the liquid fraction is used as fertiliser by spreading, compared to spreading raw manure (Beline et al., 2004; Møller et al., 2007; Popovic et al., 2012). The solid fraction can then be exported and used as organic fertiliser, usually after further treatment (drying or composting).

- Methanisation

As several nutrients such as N, P, K, Zn, Cu are retained during anaerobic digestion, the resulting digestate can be used as fertiliser (Marcato, 2007; Marcato et al., 2009; Amaral et al., 2014). However, the transformation of part of the OM into biogas leads to an increase in Zn and Cu contents in relation to the DM and an effect on their speciation (see Box 1) which is important to consider. Furthermore, Zn and Cu can also stimulate or inhibit the bacteria responsible for anaerobic digestion (Matheri et al., 2016), thus modifying biogas production.

- Drying by incineration

After incineration of the solid fraction of slurry or digestate from anaerobic digestion, two types of ash are obtained, a bottom ash (which tends to be richer and to accumulate Cu) and a so-called "fly ash" (Møller et al., 2007).

- Composting

Composting leads to the production of humic acid, the main constituent of humus, and Cu has a better affinity for this humic acid than Zn, which influences their distribution (He et al., 2009) in the final product of composting: compost. Hsu and Lo (2001) observed an increase in the soluble Cu content during the first 18 days of the composting process (up to 16% of Cu in this form) and then a decrease in this content to 3% of Cu in soluble form in the final compost. A lower effect was observed on the soluble part of Zn, which reached a maximum of 2%. Composting therefore seems to have a significant effect on the risk of Cu leaching, but less on that of Zn. Hsu and Lo (2001) observed that most of the Zn and Cu were in the low bioavailable fraction of the compost. The addition of material, such as straw or other organic substrates, or mineral additives, reduces the availability of heavy metals during composting and facilitates their transfer to more stable forms. This is related to the formation of compounds with affinity to the humic matter of the compost (Santos et al., 2018; Li et al., 2019). Furthermore, the addition of OM is also accompanied by a reduction in Zn and Cu content through dilution.

One of the purposes of these different treatments is the production of organic fertilisers or soil amendments (see Box 1) that are exported outside the livestock farms to crop systems where OM is lacking, such as market gardening. This reduces the risk of accumulation of elements such as Zn and Cu in pig farm soils by optimising their use. However, this requires that these products be standardised and consider the impact of these treatments on the speciation of the TM and therefore their bioavailability to plants.

3. Pig manure application and consequences on soils

3.4. Specific case of the return to the soil of products from the treatment of pig manure

As mentioned earlier, the treatment of pig manure does not remove Zn and Cu but concentrates them in some fractions. Møller et al (2007) indicate that application of the solid phase from centrifugal phase separation results in a lower soil load of Zn and Cu than pure slurry, but this is probably very dependent on the generally N-based fertilisation rules. Studies evaluate the germination index of seeds after fertilisation with compost, which allows to determine the phytotoxicity and maturity of the latter. According to He et al (2009), this index increases during the composting process: this means that this type of treatment leads to a decrease in phytotoxicity and improves root growth. Furthermore, this germination index can be predicted by the total content of Zn and Cu and other metals (lead).

Figure 7. Copper (Cu) and zinc (Zn) flows through the pig industry and its effluent management systems. Example of fattening pigs. Circular bioeconomy approach.

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Conclusion

Zn and Cu are essential TM for pigs but potentially toxic for the environment, and they are limited resources. It is therefore essential to characterise their flow throughout the pig production chain in order to limit their use and application to agricultural land. Figure 7 summarises this feed-animal-manure -treatment-soil continuum in the context of the circular bioeconomy.

Reducing dietary Zn and Cu, while respecting the animal's requirement, is the main way to reduce the content of these two elements in pig excrement. This can be achieved by adapting inputs as closely as possible to changing needs and by using more bioavailable sources. Effluent treatment then allows better management of the return of Zn and Cu to the soil by modifying their speciation and their content in relation to the DM. Separative treatments allow a redistribution of these elements between two phases and facilitate their export to regions with soils that are less rich in Cu and Zn, or even deficient, and on crops that have a high need.

Authors' contributions

Emma Gourlez, Fabrice Beline, Jean-Yves Dourmad, Alessandra Monteiro, Francine de Quelen participated in the definition and content of the article. Emma Gourlez wrote the manuscript and considered the corrections made by the other authors. Francine de Quelen is the coordinator of the project funding this study, which was carried out as part of a thesis directed by Fabrice Beline. All co-authors have read and approved the final version of the article.

Acknowledgements

This synthesis was conducted as part of the RECUIZ project "Optimisation of copper and zinc recycling, from their use in animal feed to their return to the soil", financed by the ADEME's GRAINE call for projects and Emma Gourlez's CIFRE thesis co-financed by Animine.

This article was first translated with www.DeepL.com/Translator, and reviewed by the authors.

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