Physico-chemical properties
The important soil property that determines acidity or alkalinity is pH and it is considered as critical parameter for healthy plant growth. During the present study soil samples studied were found to be slightly alkaline (7.92–8.38). Vanita et al. (2014) observed that agricultural soil samples from Amritsar were slightly alkaline with pH ranging from 7.4 to 7.8. Water holding capacity of soil samples ranged from 35.57–36.32% during present study. The content of calcium (mg/g) in agricultural soil samples varied from 0.80 (BLE: Baloke) to 1.63 (HBN: Hambran). All the soil samples showed magnesium content above the safe limits of 0–500 mg/kg as presented by Awashthi (1999), Indian Standard Institution (1983) and Alghobar and Suresha (2017) except for the Hambran (HBN) site which has magnesium content as (0.44 mg/g).
Potassium contents was observed to be low in comparison to calcium and magnesium. It might be due to the potential of monovalent cations (K+ and Na+) leaching from the system while divalent cations (Ca2+ and Mg2+) can be strongly adsorbed to soil particles when soil moisture content rises during rice cultivation. Application of magnesium fertilizers such as magnesium nitrate and magnesium sulfate could be the cause of high magnesium levels in the sites studied. Similar results were reported earlier by Kaur et al. (2022) in roadside soil samples collected from the vicinity of Buddha Nullah, Ludhiana, Punjab where the contents of calcium and magnesium were found in the range of 0.12–3.13 mg/g and 0.032–0.609 mg/g. Concentration of sodium in all soil samples ranged from 0.45 mg/g (BLE) to 0.62 mg/g (AWL). High content of Na in soil might be due to runoff of pesticides, fertilizers and other soil amendments. Electrical conductivity (EC) of the soil is a key indicator of its health. EC was found in the range of 23–35 mS/cm which was higher than the permissible limit i.e. 4 mS/cm (Alloway 1990). EC was observed to cause salinity issue in agricultural soil over the years (Bashir et al. 2019). The quantity of accessible nitrogen in the soil is measured in the form of nitrate-nitrogen (NO3-N) and its concentration varies from crop to crop. However in general, a preferred concentration range of 0.01–0.05 mg/g is required. High level of nitrates can form carcinogens and can accelerate eutrophication in surface waters. The levels of NO3-N can vary greatly with fluctuation of soil water. In the present study, nitrate content ranged from 0.072–0.075 mg/g. In the soil samples, phosphorous as phosphate was observed to be in the range of 0.029–0.070 mg/g where optimal phosphorous levels stimulate robust root and shoot development (Lambers 2022).
Heavy metal contents in soil
The concentration of essential micronutrient such as zinc in all the soil samples was found to be within permissible limits while Cd, Cr, Co, Cu and Pb exceeded the permissible limits. Extensive use of pesticides and fertilizers all over the countries, could pose severe risk of heavy metal contamination of soil. The occurrence of heavy metals in soil samples during present study can be correlated to the same along with the used irrigated water from Bhuddha Nullha, flowing nearby.
The high amount of heavy metal in soil samples were also reported by Kaur et al. (2014) in agricultural soils of Punjab where Cadmium (Cd) was found much higher (9.70–30.0 mg/kg) as compared to its safe limits (3–6 mg/kg) required for any agricultural soil as given by Awashthi (1999) and by Dheri et al. (2007) in agricultural soil samples of other parts of Punjab. Chromium (Cr) is a potentially toxic element (PTE) that is found mostly in areas of high human activity, such as steel and alloy manufacture, leather tanning, electroplating, wood preservation, printing and dyeing, chemical production, among others and is bound to influence plants and animals (Yang et al. 2021). It is the most commonly found in trivalent (+ 3, chromic) or hexavalent (+ 6, chromate) forms. Potassium chromate and potassium dichromate are two commercially accessible forms of hexavalent chromium (Cr(VI)). Potassium chromate is used as a fungicide to kill insects and protect plants against fungal infestations in agriculture (Gavris et al. 2014). Chromium causes the degradation of photosynthetic pigments in plants which interrupts the photosynthetic process. High level of hexavalent chromium (40 ppm − 500 ppm) in soil reduced seed germination upto 48% in bush bean Phaseolus vulgaris and by 23% in Lucerne (Medicago sativa) (Parr et al. 1982; Peralta et al. 2001). The concentration of chromium in the agricultural soil samples in the study area varied from 1.55–115.75 mg/kg and was less than the safe limits of 100 mg/kg as given by Agarwal (2009), except at one site i.e. Hambran (HBN) where the value was 115.75 mg/kg. The main reason for the high chromium content in the study area could be attributed to various anthropogenic actions, i.e., waste disposal from different industrial units, along with sampling sites in the study area.
Among the heavy metals, Cd is regarded as a serious toxicological concern (Dhaliwal et al. 2020). The maximum concentration of Cd (3.35 mg/kg) was observed at site Malakhpur (MKR) while minimum concentration of Cd (1.61 mg/kg) at site Aliwal (AWL). The use of water from the nearby flowing polluted Bhuddha Nullah, for irrigation purposes could be responsible for the high levels of Cd in the study locations exceeding the permissible limit of 0.06 mg/kg Agarwal (2009). The findings are comparable to those of Kaur et al. (2022) where Cd content (0.03 to 0.46 mg/kg) in most of the roadside samples were above the safe limit. Farid et al. (2015) reported that Cd enters the soil through the use of phosphate fertilizers, Cd-contaminated sewage sludge and manure with phosphate fertilizers, being the primary source of Cd pollution, in agricultural soil. In the current study, Pb contents in soil samples HBN (14 mg/kg) and MKR (31.5 mg/kg) were found to be above than the typical soil range i.e 10 mg/kg and below the typical range in sample AWL (2.66 mg/kg) and BLE (3.5 mg/kg) given by Agarwal (2009). Liang et al. (2011) had also reported the higher Pb concentration (44.9 mg/kg) in soil in relation to the present study. Rubber industries in the area could be accountable for the elevated Pb levels in the area. The rubber automotive tire usage was considered to be the source of contamination of soil which released manganese, iron, cobalt, nickel, copper, zinc, cadmium and lead which are used in the tire tread (Staszak 2018). Lead frequently gets used in the production of paints, putties, and pesticides as well as in arsenical pesticides such as lead arsenate (Wuana et al. 2011).
Cobalt (Co) is a component of vitamin B12, and is beneficial to humans, yet excessive amounts of cobalt can be harmful to human health (Mahey et al. 2020). The concentration (mg/kg) of cobalt in the agricultural soil samples in the study area varied from 6.35–10.16 and was less than the safe limit of 8 mg/kg (Kaur et al. 2022) except for one site, i.e., Hambran (HBN) where the value was 10.16 mg/kg. The main sources of cobalt contamination in the environment have been identified as rechargeable lithium-ion batteries, compounds containing cobalt sewage effluents, and urban and agricultural runoff (Juraszek et al. 2020). Cobalt toxicity reduces plant growth, biomass, and chlorophyll content by interfering with antioxidant enzymatic activities, photosynthetic pigments and nutrient status (Salam et al. 2022). Copper (Cu) in soils exists in different forms, and its availability to plants varies substantially. Copper is considered as a micronutrient for plants, although it is also potentially toxic. Xiong et al. (2006) observed shorter root length, fewer leaves, negative impact on nitrogen metabolism and decrease in plant biomass with increasing Cu concentrations in Chinese cabbage. Main sources of Cu in agricultural soil were reported to be application of biocides or fungicides (Liscakova et al. 2022). In this study, the Cu concentrations were found 39.45 mg/kg in Hambran and 30.75 mg/kg in Malakhpur which were above the maximum permissible limits (20 mg/kg) as given by Agarwal (2009).
During the present study at all study sites, the content of zinc was observed to be less than the maximum permissible limit of 50 mg/kg (Agarwal 2009). Similar observations were also made by Chen et al. (2022). Zinc is a micronutrient that both plants and animals require. Zn deficiency results in yellowing of leaf, reduced plant growth and cause senescence (Kumar et al. 2022; Asati et al. 2016). Zn enters the environment through a variety of pathways, including mining, smelting, industrial and municipal wastes, urban runoff and most notably erosion of Zn-containing soil particles (Noulas et al. 2018). Nickel (Ni) is a micronutrient that is required for plant development and for the activity of several enzymes in low concentrations and is toxic at high concentrations. The presence of Ni in soil can be attributed to mining operations, smelter emissions, coal and oil combustion, sewage, phosphate fertilizers and pesticides (Asati et al. 2016). The critical toxicity limit of Ni for sensitive, moderately sensitive and resistant plants was suggested to be greater than 10 mg/kg, 50 mg/kg, and 1000 mg/kg dry matter, respectively (Dehdezi et al. 2021). Wuana et al. (2011) observed that the connection between soil and water contamination and metal uptake by plants is influenced by a variety of chemical and physical soil factors as well as crop physiological properties. Setia et al. (2021) reported diffuse sources of metals in soil, to be fertilizers, pesticide, sediments runoffs and higher concentration of metals in soil along trans-boundary areas of Sutlej river (Punjab). Tariq and Rashid (2012) reported that nitrate fertilizers were the most significant sources of Cd and Co while fungicides and pesticides for Pb and fertilizers for Zn, Cu, Fe, and Mn (Kaur et al. 2022). Furthermore, the sources of heavy metals like Cd, Pb and Co in agricultural soil samples, in the present study may be mainly originated from pesticides, fertilizers and industrial or domesticated sewage as well as irrigated water from Bhuddha Nullha.
Genotoxic potential
Chromosomal abnormalities are termed as the clear indication of DNA damage that cannot be easily repaired (Zeyad et al. 2019). Since the Allium cepa is a eukaryotic system, it can provide a higher degree of proximity when compared to the expected impacts on biota exposed to harmful substances such as heavy metals, pesticides and compounds discharged from industries like dyeing, electroplating, textiles etc (Batista et al. 2016; Mazzeo et al. 2011). Changes in chromosomal structure or chromosomal number are examples of chromosomal aberrations. Breaks in DNA, suppression of DNA synthesis and changes in DNA replication can all cause changes in chromosomal structure. Various chromosomal aberrations, such as chromosomal breaks, bridges and ring chromosomes indicate clastogenic aberrations (Sabeen et al. 2020). During DNA replication clastogenic abnormalities can occur. Physiological abnormalities caused by spindle anomalies include c-mitosis, vagrant, stickiness, delayed anaphase, and laggard. Different cytological parameters such as mitotic index and chromosome abnormalities such as breaks in chromosomes, lagging chromosome, c-mitosis, bridge formation, and stickiness were used to determine the genotoxicity of agricultural soil samples.
The induction of toxicity in soil samples by lowering the mitotic index and increase in chromosomal aberration was observed in all studied soil samples in the A. cepa test. In present study, mitotic index was found least for sample Hambran (HBN: 10.8%) having Cr more than permissible limit reported by Agarwal (2009). Singh et al. (2015) also reported significant decrease in mitotic index and in root length of Allium cepa as the concentration of Cr during the experiment increased. Metal stress causes disruptions in the cell cycle or chromatin disfunction which results in a significant reduction in mitotic index (Kopliku et al. 2013). Patnaik et al. (2013) observed that Cr (VI) at higher concentrations (100 or 200 µM) was toxic and resulted in nuclear disintegration or pycnosis in root cells of Allium cepa. The genotoxic effect can also be observed through the significant increase in the frequency of chromosomal aberrations for all the soil samples. Amongst all the agricultural soil samples, HBN (Hambran) sample showed maximum (46.7%) whilst sample BLE (Baloke) showed minimum (31%) percentage of chromosomal aberrations. The treatment with control (acid-washed sand) showed 2.1% total chromosomal aberration. The frequency of cells with delayed anaphase was found to be highly elevated followed by chromosomal bridge formation, c-mitosis, abnormal metaphase and stickiness. When the two anaphasic chromosomal groups are close to each other near the equatorial plate, delayed anaphase occurs. The loss of the protein covering that shields the DNA in chromosomes, as well as the breakage and exchange of basic folding fibre units in chromatids, can cause chromosome stickiness (Kumar et al. 2021). Disruption of spindle fibre lead to the occurrence of c-mitosis during the mitotic phase (Seth et al. 2008). The presence of harmful substances in the growth media of plants could be indicated by C-mitosis (Bonciu et al. 2018). Chromosomal bridges could be formed as a result of chromosome adhesion. Which was a sign of a toxic consequence and was considered irreversible (Marcano et al. 2004; Kumari et al. 2009). Similar findings were made by Soodan et al. (2015) and Becaro et al. (2017).
The usage of a wide range of organic and inorganic pesticides, herbicides and fertilizers by the farmers, in the current study, might have contributed to the soil samples genotoxicity. Insecticides are a class of chemical compounds that have a wide spectrum of toxicity and as a result, pose a risk to the environment (Anjum and Malik 2013). Kalefetoglu Macar (2021) reported cytotoxic and genotoxic effects such as decline in mitotic index of pesticide (abamectin) in non-targeted organisms following Allium cepa root chromosomal assay. Heavy metals, pesticidies and a variety of other pollutants present in soil were responsible for decline in mitotic index and ultimately cell death in roots of A. cepa (Cortes-Eslava et al. 2018; Carita and Marin-Morales 2008). Apart from this, high genotoxicity could possibly be attributable to the application of waste water to agricultural land from Bhuddha Nullah, which is located near the test sites under investigation. Our findings are consistent with some previous study of Pohren et al. (2013) who evaluated genotoxic potential of soil contaminated with heavy metals and reported different chromosomal abnormalities following Allium cepa root chromosomal aberration assay. Other studies that revealed cytotoxicity and genotoxicity of metal like chromium in plant cells also supported the current findings (Panda et al. 2002). According to Kwankua et al. (2012), heavy metal exposure hindered cells from entering cell division stages, resulting in a decrease in mitotic index.
Statistical analysis
Statistical analysis was performed using SPSS Statistics software (IBM SPSS Statistis for window, version 21). One-way ANOVA was applied to find significant differences between values of heavy metals and genotoxicity parameters like mitotic index (MI) and total aberrant cells (TAC %) for collected agricultural soil samples. One-way ANOVA and post hoc analysis using the Tukey test have showen significant differences in the observed parameters. Pearson correlation analysis was carried out using different agricultural soil quality parameters which revealed that the content of Cd, Cr, Cu, Co, Ni, Pb and Zn were strongly affected by pH. Soil pH is inversley correlated with these metals and the highest correlation coefficient was found for Zn (r = − 0.889). Significant positive correlations were observed betweeen Cd and Pb in soil samples (r = 0.981, p ≤ 0.01), which suggested the similar pollution sources of these two heavy metals. Similarly, significant positive correlation were observed between Cu and Co (r = 0.989, p ≤ 0.01) and between Cu and Ni (r = 0.981, p < 0.05) and between Cu and Zn (r = 0.973, p < 0.05). These strong positive correlation between these heavy metals, suggested the similar origin of these metal pairs probably from agrochemicals used in the farm. Ihedioha et al. (2016) have reported strong positive and significant correlation between Cr, Ni and Mn in soil from Ada field, Nigeria and has implied that the metals had the same pollution sources. Mitotic index exhibited negative correlation with total aberrant cells. Cu, Co and Ni showed significant negative correlation with mitotic index, and significant positive correlation with TAC, indicating genotoxic nature of these metals.