Strawberry is a very attractive crop producing fruits with excellent organoleptic characteristics coupled with positive nutritional and nutraceutical attributes. In Mediterranean countries, strawberry production is around 1.6 million tons annually, comprising almost 18% of the world production (Chandler et al., 2012; FAOSTAT 2020). Mediterranean area suffer for water scarcity and climatic changes will exacerbate this situation (IPCC, 2021). Since water demand in strawberry is very high, alternative source of water like wastewater will be precious (Pedrero et al., 2018; Renai et al., 2021). Strawberry is normally produced in protected cultivation or in open field. Open field may be very susceptible to Cd contamination with damage to plants (Zhang et al., 2020). Among the negative effects of Cd stress there are a reduction on the content of soluble sugar and strawberry yield (Zhang et al., 2020). In our experimental condition, Cd reduced the strawberry weight of 50% compared to control strawberries. Our data showed that the plants treated with Cd had significant decrease of Chl a + b compared to control after 28 days. Cadmium toxicity effects on chlorophylls could be explained because of inhibitory effect of Cd on enzymes involved in pigment biosynthesis (Muradoğlu et al., 2015). Furthermore, Cd induced oxidative damage and usually leads to the degradation of pigments (Chen et al., 2003). Cadmium has been shown to interact with the uptake of numerous elements (Haider et al., 2021). In sugar beet, deficiency of Fe is induced by Cd in roots (Chang et al., 2003). The uptake of Ca, Zn, Mn in pea was strongly inhibited after exposure to Cd (Metwally et al., 2005). In saltbush (Atriplex halimus L.), a decreased uptake of Ca was found due to the toxicity of Cd (Kinay, 2018). In strawberry Camarosa cultivar (Muradoğlu et al., 2015), increasing Cd affect other mineral elements. Zn concentration was found higher in Camarosa root when Cd concentrations increase. An opposite trend has been observed in our experiment, where Zn decrease in root under Cd exposure. It was also demonstrated that concentration of Fe, Ca, and Zn in strawberry leaves decreased with the increase of Cd concentration (Bi-qing et al., 2007). In our study, Ca and Zn concentration in strawberries showed a significative interaction between Cd and glufosinate with a lesser concentration of these in treated plants compared to control.
Glufosinate is considered a contact herbicide due to its fast activity and limited translocation in plants. The absence of active transporters and physicochemical characteristics that facilitate its translocation are the main reasons of its low translocation rates (Takano et al., 2020a). An efficient uptake of glufosinate depends on several factors, such as, spraying conditions, air temperature, humidity, and the target species. Low temperature and low humidity reduce glufosinate uptake (Takano et al., 2020). Our data demonstrated that glufosinate has been uptaken by strawberry and degraded to MPP metabolite. Glufosinate uptake and degradation did not affect plant growth and fruits quality.
The use of glufosinate ammonium causes glutamine synthetase inhibition with ammonium ion accumulation and alteration of photosynthesis. A previous study (Petrovic et al., 2009) investigated the relationship between excess NH4-N in foliar tissues and interveinal chlorosis in leaves of “Nyoho” strawberry, a cultivar that often suffers leaf-yellowing symptoms. Glufosinate dose of 185 mg L–1 was shown to be lethal to “Nyoho”, inducing severe interveinal chlorosis that progressed into necrosis and rapid desiccation of the leaf tips. In accordance with this study, we observed a significative reduction in chlorophyll a + b concentration in plants treated with glufosinate ammonium.
Most abiotic stress conditions like heavy metal toxicity result in increased production of the plant hormone ethylene (Maksymiec, 2007). Ethylene synthesis increased after inhibition of the photosynthetic apparatus by heavy metals, probably resulted from increased activity of enzymes synthesizing this hormone after heavy metal action. In barley plants, Vassilev et al. (2004) showed that increasing Cd concentrations (0, 14, 28, and 42 mg kg− 1), significantly increased ethylene production by 29 and 44% at the 14 and 28 mg kg− 1 and significantly decreased it by 29% at the 42 mg kg− 1 after 10 days of treatment. Our observation about ethylene did not show any significant difference in the two sampling between Cd and control treatments, but we used realistic Cd concentrations of 1 mg L− 1.
Ethylene evolution is also an indicator of herbicidal action. You and Barker (2002) showed that ethylene evolution rose with increase of glufosinate ammonium concentration to 25 mg L− 1 in tomato plants. Despite this evidence, the ethylene measurement did not show any significant difference in our experiment. Again, these evidences in our study showed that realistic residual concentrations of glufosinate (10 µg L− 1) in the wastewater have no impact on ethylene production.
Although heavy metals and pesticides are individually toxic, the interaction between the two should be evaluated. This relation can be synergistic or antagonistic. The synergism occurs when the joint toxicity increase (compared to the individual one) while the antagonism happens when the joint toxicity decrease (Alengebawy et al., 2021). A study conducted by Wang et al. (2015) presented the effect of combined toxicity of Cd with five types of insecticides on earthworm. Eleven mixtures had synergistic effects while 5 exhibited antagonistic ones: this evidence showed that synergic effects occurred more than antagonistic one. To date, there is little data on the effects of joint toxicity in plants. Examples about the interaction between heavy metals and pesticides in plants regarded the use of Cd and Acetochlor or Bensulfuron-methyl as pesticides in rice seedlings (Oryza sativa L., cv. Jinyou 402). In these experiments, treated plants showed a synergic interaction decreasing soluble protein content and suppressing roots and shoots growth (Huang and Xiong, 2009). In our experiments, we observe an antagonistic effect, with a decrease in the joint toxicity for the chlorophyll a + b concentration.
As reported in the Regulation (EC) n. 1881/2006 (OJ L 364 2006), human foodstuffs have been regulated for the maximum levels of some heavy metals. As concern Cd, the maximum acceptable concentration in strawberry was set to 0.050 mg kg− 1 wet weight. The 57% of examined strawberry grown under Cd exposure exceed the Cd concentration indicated by EU and the 100% the fruits grown under the mix Cd and glufosinate. Following the public health literature information about Acceptable Daily Intake of Cd (Melai et al., 2018), our results about EDI calculated on the base of maximum concentration measured of Cd (worst-case) and the more exposed consumers (people from 10 to 18 years) is not ever exceeded the indicated value. Since in literature the data of MPP accumulation in human body are not available, we can only report that the intake of strawberry fruits growth under our experimental condition present this molecule.
In conclusion, this work proves that realistic and residual concentrations of Cd and glufosinate ammonium, that could be present in wastewater, have an impact on strawberry growth and fruit production. However, although the quality of strawberry fruits did not show noticeable change in terms of Brix and polyphenols, it has been demonstrated that in hydroponic conditions Cd and MPP metabolite can be present in the fruits representing a potential risk to human health. Anyway, considering that the same waters can also be used in open fields where the soil plays a key role in the retention of pollutants, the toxicity is even less high. The use of wastewater like alternative water source for agriculture fits into the circular economy (CE) concept in which waste should be treated as a secondary raw that can be recycled to be reused. Our data confirmed the importance of understanding how contaminants interact each other and translocate in edible parts like fruit. Only this approach will make wastewater use possible and safe for human, plants, and the environment.