Zletovska River Research Articles

Pollution impact on metal and biomarker responses in intestinal cytosol of freshwater fish.

In the present study, essential and nonessential metal content and biomarker responses were investigated in the intestine of fish collected from the areas polluted by mining. Our objective was to determine metal and biomarker levels in tissue responsible for dietary intake, which is rarely studied in water pollution research. The study was conducted in the Bregalnica River, reference location, and in the Zletovska and Kriva Rivers (the Republic of North Macedonia), which are directly influenced by the active mines Zletovo and Toranica, respectively. Biological responses were analyzed in Vardar chub (Squalius vardarensis; Karaman, 1928), using for the first time intestinal cytosol as a potentially toxic cell fraction, since metal sensitivity is mostly associated with cytosol. Cytosolic metal levels were higher in fish under the influence of mining (Tl, Li, Cs, Mo, Sr, Cd, Rb, and Cu in the Zletovska River and Cr, Pb, and Se in the Kriva River compared to the Bregalnica River in both seasons). The same trend was evident for total proteins, biomarkers of general stress, and metallothioneins, biomarkers of metal exposure, indicating cellular disturbances in the intestine, the primary site of dietary metal uptake. The association of cytosolic Cu and Cd at all locations pointed to similar pathways and homeostasis of these metallothionein-binding metals. Comparison with other indicator tissues showed that metal concentrations were higher in the intestine of fish from mining-affected areas than in the liver and gills. In general, these results indicated the importance of dietary metal pathways, and cytosolic metal fraction in assessing pollution impacts in freshwater ecosystems.

Accumulation and distribution of heavy metals in soil and food crops in the Pb–Zn mine environ. Case study: Region of Probištip, North Macedonia

The aim of the study was to investigate the exploration and distribution of various chemical elements in soils and vegetables from gardens in the region of Probištip, North Macedonia. This region was exposed to heavy metals pollution from Pb–Zn mining and flotation tailings landfill near the town of Probištip for 70 years. A total of 38 samples of various crops and vegetable products from 19 gardens were analyzed. The analyzes were carried out by ICP-AES (Al, Ba, Ca, Cr, Cu, Fe, K, Li, Mg, Mn, Na, Ni, P, Pb, Sr, V, and Zn) and by ICP-MS (As, Cd, Co, Cr, and Pb). It was found that the content of some potentially toxic elements (As, Cd, Cu, Pb, and Zn) in the soil samples from the gardens, exceeded the target values. From these results, it is clear that the contamination of the soils in this area is due to mining activities and the distribution of flotation wastes from the nearby flotation dump. It was also found that the contaminated soils lead to elevated levels of these elements in some vegetables from the gardens, especially those near the Zletovska River and the flotation waste landfill, exceeding the nationally permissible food content.

Effects of water contamination with heavy metals on pigmented macrophages in spleen of Vardar chub (Squalius vardarensis Karaman) from three rivers in the North-Eastern region of North Macedonia

Background and purpose: Pigmented macrophage aggregates (PMAs) are a part of the immune system of fish and are considered as valuable biomarkers of the impact of water pollution on fish health.Materials and methods: Immunomodulatory effect of water contaminated with high levels of metals on spleen and splenic PMAs of Vardar chub (Squalius vardarensis) (n=129) was examined in two seasons, spring, and autumn 2012. For this purpose, three rivers were chosen: the Zletovska River, heavily contaminated with metals, the Kriva River, moderately contaminated with metals combined with high level of organic matter, and the Bregalnica River, without heavy metal contamination. Spleen masses and splenosomatic indices (SSI) were used for determining the general impact of water contamination on the immune system of Vardar chub. Moreover, volumes and numbers of PMAs within spleen tissue were determined stereologically.Results: The isolated effect of high exposure to metals caused the reduction of spleen mass and/or SSI in the fish from the Zletovska River. The combined effect of exposure to metals and high levels of nutrients in the fish from the Kriva River referred to stimulation of PMAs response, namely increased volumes of splenic PMAs. The existence of seasonally dependent differences was noted since the observed variability in PMAs between rivers occurred only in the autumn.Conclusions: Splenic PMAs in the Vardar chub can be used as an additional biomarker of metal pollution, but with caution, since enhancement of the volumes and numbers of MACs depends on the exposure level of heavy metals, on the presence of the other contaminants, as well as on the physiological condition of the fish.

The effect of different pollutants exposure on the pigment content of pigmented macrophage aggregates in the spleen of Vardar chub (Squalius vardarensis Karaman, 1928).

Pigmented macrophage aggregates (MAs) are known to change under influence of various factors, such as aging, season, starvation, and/or pollution. In this study, changes in the pigment content of the MAs in the spleen of Vardar chub (Squalius vardarensis, Karaman) (n = 129) collected in spring and autumn, from three rivers with different pollution impact was examined: Zletovska River (metals), Kriva River (metals and municipal wastewater), and Bregalnica River (municipal wastewater). Collected data revealed increased relative volume and number of MAs containing hemosiderin under the influence of metals, significant in autumn (p < .05). In chub exposed to metals combined with municipal wastewater, significant increase of lipochrome accumulation in MAs in autumn, melanin in MAs in fish captured in the spring season, and number of splenic MAs containing combination of melanin and lipochrome was noted. Volumes and number of MAs containing combination of hemosiderin and lipochrome increased in spleen of fish captured in autumn from both Zletovska River and Kriva River, most likely due to the contribution of hemosiderin and lipochrome, respectively. Values measured for the various pigments in splenic MAs in fish captured from Bregalnica River, were overall closer to the values measured for fish captured from Kriva River. Notably, melanin and lipochrome are more likely to be found in fish from waters influenced by municipal wastewater (organic pollution) and hemosiderin in fish spleen from water influenced by mining activity (heavy metals pollution).

Mining waste as a cause of increased bioaccumulation of highly toxic metals in liver and gills of Vardar chub (Squalius vardarensis Karaman, 1928)

Freshwater contamination with mining waste can result with high concentrations of toxic metals in the water and in fish organs. In North-Eastern Macedonia, several rivers (e.g., Zletovska, Kriva) are exposed to acid mine drainage from active Pb/Zn mines. Previous studies confirmed high concentrations of dissolved metals in their water. This study was performed in liver and gills of Vardar chub (Squalius vardarensis Karaman, 1928) from three Macedonian rivers (Bregalnica, Kriva and Zletovska) in spring and autumn 2012. The aim was to establish if increased exposure to certain metals have resulted with their increased bioaccumulation. The concentrations of 19 elements were measured in cytosolic tissue fractions, to obtain information on metabolically available metal species. The following ranges of cytosolic concentrations of highly toxic elements were measured in the Vardar chub liver (in μg/L): Cd, 1.18–184; Cs, 0.25–25.4; Tl, 0.02–5.80; Pb, 0.70–61.1. Their ranges measured in the gills (in μg/L) were the following: Cd, 0.24–59.2; Cs, 0.39–24.4; Tl, 0.01–1.00; Pb, 0.65–87.2. Although the water of the mining impacted Zletovska River was highly contaminated with several essential metals, especially with Mn and Zn, the majority of essential elements (Na, K, Ca, Mg, Co, Cu, Fe, Mn, Mo, and Zn) did not reflect the exposure level. In contrast, seven nonessential elements reflected the level of exposure in the water. Significantly increased hepatic and gill concentrations of Cs, Rb, Sr, and Tl were detected in Vardar chub from the Zletovska River compared to the other two rivers, of Cd and Pb in the Zletovska and Kriva River compared to Bregalnica, and of V in the Bregalnica River compared to Zletovska and Kriva rivers. Observed significant metal bioaccumulation, in particular of highly toxic elements, as a consequence of exposure to water contaminated with mining waste points to necessity of intensified supervision of mining impacted rivers.

Malondialdehyde concentrations in the intestine and gills of Vardar chub (Squalius vardarensis Karaman) as indicator of lipid peroxidation.

A lipid peroxidation product, malondialdehyde (MDA), was studied in Vardar chub (Squalius vardarensis Karaman) as an indicator of oxidative stress, using native fish from three rivers in northern Macedonia: the mining-impacted Zletovska and Kriva rivers and the agriculturally impacted Bregalnica River. MDA concentrations were measured in the intestine in the spring and autumn of 2012 and in the gills in autumn. The aims of the study were to establish the type of contamination which provokes a more pronounced MDA increase, as well as the organ which more reliably reflects the occurrence of oxidative stress. MDA levels in the intestine in spring amounted to 3.29-155.8nmolg-1 and in autumn to 4.85-111.1nmolg-1, whereas MDA concentrations in the gills in autumn were 7.69-147.5nmolg-1. Stronger influence of organic contamination on development of oxidative stress was observed in both organs, as seen from higher median MDA concentrations in autumn in fish from the highly pesticide-contaminated Bregalnica River (gills 78.4nmolg-1; intestine 23.5nmolg-1) compared to the highly metal-contaminated Zletovska River (gills 15.9nmolg-1; intestine 17.4nmolg-1). The response of the gills to contamination was twice stronger than that of the intestine. The majority of fish from the pesticide-polluted river had increased MDA in the gills, in contrast to only sporadically increased MDA in the intestine. Our results indicated that development of oxidative stress strongly depends on the selected fish organ and that the gills seem to be a better choice for monitoring oxidative stress than the intestine, due to their continuous and direct exposure to polluted river water.

Effects of heavy metal pollution on pigmented macrophages in kidney of Vardar chub (Squalius vardarensis Karaman).

Pollution with heavy metals may influence the immune system of fish, leading to impairment of their health or even increase their mortality. The fish kidney is one of the first fish organs to be affected by water contamination. Amounts of kidney macrophages (MACs), which are involved in fish immune response, as well as the qualitative and quantitative changes in the pigmented MACs in fish kidney, are used as biomarkers of pollution. Therefore, in this study, we have evaluated relative and total volumes of trunk kidney pigmented MACs, and analyzed the pigments accumulated within them. Fish were sampled from two mining impacted rivers, Kriva and Zletovska, highly contaminated with heavy metals, and from one reference river, Bregalnica, in spring and autumn of 2012. We have observed that main pigments found in kidney MACs of Vardar chub were melanin and lipofuscin/ceroid, as well as that relative volumes of MACs ranged from 0.56 to 1.68%. Moreover, the results showed that relative volumes of pigmented MACs were higher in metal contaminated rivers, especially in autumn season in the Zletovska River, concurrently with extremely high metal exposure. In addition, condition factors and kidney somatic indices were found significantly lower in the Zletovska River in both seasons, autumn and spring, possibly also as a consequence of high water pollution. Our data confirm that increase in relative volumes of pigmented MACs may serve as warning sign of potential heavy metal pollution in aquatic environment.

Broad spectrum screening of 463 organic contaminants in rivers in Macedonia

Target screening of 463 organic contaminants in surface water using ultra high performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) with direct injection was performed in spring of 2015 in northern Macedonia, at six sampling sites in four rivers belonging to Vardar basin: Kriva, Zletovska, Bregalnica and Vardar. The aim of the study was to differentiate between various types of organic contamination characteristic for different types of anthropogenic activities, such as mining, agriculture, and urbanization. Depending on the studied river, 9–16% of analyzed compounds were detected. The highest total levels of organic contaminants were recorded in agriculturally impacted Bregalnica River (1839–1962ngL−1) and Vardar River downstream from the city of Skopje (1945ngL−1), whereas the lowest level was found in the mining impacted Zletovska River (989ngL−1). The principal organic contaminants of the Bregalnica River were herbicides (45–55% of all detected compounds; 838–1094ngL−1), with the highest concentrations of bentazone (407–530ngL−1) and molinate (84–549ngL−1), common herbicides in rice cultivation. The main organic contaminants in the other rivers were drugs (70–80% of all detected compounds), with antibiotics as a predominant drug class. The highest drug concentrations were measured in the Vardar River, downstream from Skopje (1544ngL−1). Screening of surface water by UHPLC-QTOF-MS was proven as a practical tool for fast collection of comprehensive preliminary information on organic contamination of natural waters, which can present a significant contribution in the monitoring and preservation of good ecological status of freshwater ecosystems.

Histopathology investigation on the Vardar chub (Squalius vardarensis) populations captured from the rivers impacted by mining activities

Many natural freshwater ecosystems, especially in the north eastern Macedonia, are polluted with heavy metals, which are released by active mines. Long-term exposure to high levels of dissolved metals might result in increased metal bioaccumulation in organs of aquatic organisms, and consequently might cause various sub-toxic and toxic effects. The aim of this study was to assess the health of Vardar chub (Squalius vardarensis) inhabiting mining impacted rivers Zletovska and Kriva, in comparison with chub from the reference Bregalnica River. It was done by use of indicators of tissue damage (histopathology of liver and gonads) and general indicators of exposure to environmental stressors (condition factor, organo-somatic indices and external/internal macroscopic lesions). Histological assessment of gonads revealed good reproductive health in all three rivers, indicating high tolerance of gonads to contaminant exposure. Contrary, several external/internal lesions were more pronounced in chub from severely metal contaminated Zletovska River. Prevalence of hepatic lesions was also higher in mining impacted rivers (in Kriva, 70%; in Zletovska, 59%) compared to Bregalnica River (38%). The spectrum of histological lesions observed in chub liver varied from non-specific minor degenerative conditions, such as lymphocyte infiltration, fibrosis, parasites, granulomas and lipidosis, to extensive and/or more severe changes such as bile duct proliferation, necrosis, megalocytosis, light-dark hepatocytes and hepatocytes regeneration. The results of histopathological investigation for all three rivers showed clear signs of water contamination, especially prominent in mining influenced rivers. More research efforts should be devoted to study of environmental conditions and metal contamination in the mining impacted rivers worldwide, especially of their effects on health of local ichthyofauna.

Evaluation of histopathological alterations in the gills of Vardar chub (Squalius vardarensis Karaman) as an indicator of river pollution

Quantification of histopathological alterations in the gills of Vardar chub (Squalius vardarensis Karaman) was performed in 2012 in rivers of north-eastern Macedonia, with the aim to examine the effects of water quality in the rivers (Zletovska and Kriva River-impacted by active Pb/Zn mines; Bregalnica River-contaminated by agricultural waste). The biological alterations in chub were classified as: circulatory disturbances, regressive and progressive changes, but their severity differed. Altogether the mildest changes were observed in the gills of chub from the Bregalnica River, a less polluted river, whereas mining impacted rivers were characterized by more severe alterations. In the gills of chub from the Zletovska River, which is highly contaminated with numerous metals, sulphates and chlorides, the highest lesion indices were found for the regressive changes of both epithelium and supporting tissue, with typical lesions referring to atrophy, thinning and lifting of epithelial cells, necrosis of epithelium and chloride cells, as well as deformations of lamellar cartilaginous base. Gill damages of chub from the Kriva River were overall milder compared to the Zletovska River, in accordance with pollution status. In the gills of chub from that river, progressive changes were more pronounced, specifically severe hyperplasia of mucous cells and epithelium in the interlammellar space, leading to fusion of lamellae, as well as hypertrophy of chloride cells. The comparison between seasons indicated higher intensity of progressive changes in all three rivers in autumn, when water level was very low, and consequently, water contamination was more pronounced due to concentration effect. The pattern and severity of histopathological alterations in the chub gills reflected differences in contamination levels and type of contaminants in different rivers and sampling periods, and thus have been proven as a valuable indicator of water quality

Surface Water Characterization of Three Rivers in the Lead/Zinc Mining Region of Northeastern Macedonia

Mine waste is recognized as being one of the most serious threats for freshwater ecosystems, and it still represents one of the greatest environmental concerns in Macedonia. The aim of our investigation was to obtain an in-depth understanding of mining influence on freshwater systems from water contamination to effects on aquatic organisms. In this study, we assessed the impact of active lead (Pb)/zinc (Zn) mines Zletovo and Toranica on the water quality of three rivers in northeastern Macedonia (Bregalnica, Zletovska, and Kriva rivers) based on data collected in spring and autumn of 2012. The Bregalnica River, near Shtip, was characterized mainly by weak contamination with arsenic, barium, iron, molybdenum, titanium, uranium, vanadium, nitrate, and phosphate, as well as critical faecal pollution, which alltogether could be connected to agricultural activities; however, an impact of the mines was not observed. Contrary, both the Zletovska and Kriva rivers showed a clear impact of Pb/Zn mines on water quality. In the Zletovska River, increased concentrations of cadmium (Cd), cobalt, cesium, copper, lithium, manganese (Mn), nickel, rubidium, tin, strontium, thallium, Zn, sulphates, and chlorides were found, especially in autumn (e.g., Cd 2.0 μg L(-1); Mn 2.5 mg L(-1); Zn 1.5 mg L(-1)). In the Kriva River, increased Cd (0.270 μg L(-1)) and Pb (1.85 μg L(-1)) concentrations were found only in spring, possibly due to sediment resuspension during greater water discharge. The selected sampling sites on the Bregalnica, Zletovska, and Kriva rivers were confirmed as being appropriate locations for further studies of mining waste's impact on freshwater ecosystems, the first one as a nonimpacted site and the other two as possible areas of increased exposure of aquatic organisms to metals.

Heavy metal characteristics in Kočani Field plant system (Republic of Macedonia)

Heavy metal contamination of soils is widespread and induces a long-term risk to ecosystem health. This research focuses on the heavy metal contamination, transfer values and risk assessment in the Kočani Field plant system (Republic of Macedonia). To identify the heavy metal concentrations in Kočani crops (rice and maize), the geochemical analysis was performed by inductive coupled plasma mass spectrometer and thereupon the transfer factor (TF) and estimated daily intake amount values were calculated. The highest As, Cd, Mo, Pb and Zn values were determined in the rice samples grown in the paddy fields near the Zletovska River. The highest Pb and Mo concentrations measured in the maize samples were from the maize fields near the Zletovska River and Ciflik city. High TF values for Mo, Zn, Cd and Cu revealed a strong accumulation of Mo, Zn and Cd by rice and Mo and Zn by maize crops. The results of the estimated daily intake showed that the regular consumption of rice and maize crops containing the highest Cd, Mo, Pb and Zn concentrations could pose a serious threat to human health, because the daily intake of Cd, Mo, Pb and Zn for crops grown in the fields around the Zletovska River exceeded the recommended provisional tolerable daily intake values. Taking into account the results, the area around Zletovska River is considered as the most anthropogenically impacted part of Kočani Field.

Distribution and mobility of heavy metals in paddy soils of the Kočani Field in Macedonia

Contamination of soils with heavy metals is widespread and poses a long-term risk to ecosystem health. Abandoned and active mining sites contain residues from ore-processing operations that are characterised by high concentrations of heavy metals. The distribution and mobility characteristics of heavy metals (As, Cd, Cu, Pb, and Zn) in paddy soil samples from Kočani Field (Macedonia) using ICP-EAS and a sequential extraction procedure was evaluated. The results indicate that highly elevated concentrations of As, Cd, Cu, Pb, and Zn were detected in the paddy soil sample from location VII-2 in the vicinity of Zletovo mine and Zletovska river in the western part of Kočani Field, which drains the untreated acid mine waters and mine wastes from the active Zletovo mine. The degree of contamination based on index of geoaccumulation (I geo) from strong to weak in the paddy soils samples is Pb > As > Cd > Zn > Cu. The mobility potential of heavy metals in all paddy soil samples increases in the order As < Cu < Pb < Zn < Cd. According to the results of the anthropogenic impact on the paddy soils, a further study on the heavy metal concentrations in rice and other edible crops, the remediation process of the paddy soils and a dietary study of the local population are needed.

Heavy metal contamination of paddy soils and rice (Oryza sativa L.) from Kočani Field (Macedonia)

This research focuses on the heavy metal contamination of the paddy soils and rice from Kocani Field (eastern Macedonia) resulting from irrigation by riverine water impacted by past and present base-metal mining activities and acid mine drainage. Very high concentrations of As, Cd, Cu, Pb and Zn were found in the paddy soils (47.6, 6.4, 99, 983 and 1,245 microg g(-1)) and the rice (0.53, 0.31, 5.8, 0.5 and 67 microg g(-1)) in the western part of Kocani Field, close to the Zletovska River, which drains the mining facilities of the Pb-Zn mine in Zletovo. In terms of health risk, the observed highest concentrations of these elements in the rice could have an effect on human health and should be the subject of further investigations.

Major and trace elements in paddy soil contaminated by Pb–Zn mining: a case study of Kočani Field, Macedonia

The objective of this study was to assess the bulk chemical composition as well as the extent and severity of heavy metal contamination in the paddy soil of Kocani Field (eastern Macedonia). The results revealed that the paddy soil of the western part of Kocani Field is severely contaminated with Pb, Zn, As and Cd in the vicinity of the Zletovska River due to irrigation with riverine water that is severely affected by acid mine and tailing effluents from the Pb-Zn mine in Zletovo. The detected total concentrations of these metals are far above the threshold values considered to be phytotoxically excessive for surface soil. The paddy soil in the vicinity of the Zletovska River was also found to exhibit elevated levels of Ba, Th, U, V, W, Mo, Cu, Sb, Bi, Ag, Au, Hg and Tl, with concentrations above their generally accepted median concentration values obtained during this study. A correlation matrix revealed that the Mn and Fe oxides/hydroxides are the most important carrier phase for several trace elements, with the exception of rare earth elements (REEs). These also represent a major sink for the observed heavy metal pollution of the soil. REEs are mostly associated with two phases: light (L)REEs are bound to K-Al, while heavy (H)REEs are bound to Mg-bearing minerals. Although there is no direct evidence of a health risk, the paddy soil in the vicinity of Zletovska River needs further investigation and an assessment should be made of its suitability for agricultural use, particularly in view of the highly elevated concentrations of Pb, Zn, As and Cd.

Bacterial leaching of waste uranium materials

The effect of ferrobacteria and thiobacteria on the leaching of waste uranium materials from which 70-80% of uranium was previously leached by classical chemical hydrometallurgical procedure has been investigated. The bacteria used are found in the ore and the mine water of Zletovska River locality, Yugoslavia. Parameters of biological leaching were examined in the laboratory. Leaching conditions were changed with the aim of increasing the amount of uranium leached. The effect of pyrite added to the waste materials before the beginning of leaching has also been examined. Uranium leaching is directly proportional to the composition and number of ferrobacteria and thiobacteria, and increased by almost twice the value obtained from the same starting materials without using bacteria. Increased sulphuric acid concentrations stimulate considerably the rate of leaching. Uranium leaching is increased up to 20% while sulphuric acid consumption is simultaneously decreased by the addition of pyrite. Uranium concentrations in starting waste materials used for leaching were extremely low (0.0278 and 0.372% U) but about 60% recovery of uranium was obtained, with relatively low consumption of sulphuric acid.

Bacterial leaching of waste uranium materials

The effect of ferrobacteria and thiobacteria on the leaching of waste uranium materials from which 70–80% of uranium was previously leached by classical chemical hydrometallurgical procedure has been investigated. The bacteria used are found in the ore and the mine water of Zletovska River locality, Yugoslavia. Parameters of biological leaching were examined in the laboratory. Leaching conditions were changed with the aim of increasing the amount of uranium leached. The effect of pyrite added to the waste materials before the beginning of leaching has also been examined. Uranium leaching is directly proportional to the composition and number of ferrobacteria and thiobacteria, and increased by almost twice the value obtained from the same starting materials without using bacteria. Increased sulphuric acid concentrations stimulate considerably the rate of leaching. Uranium leaching is increased up to 20% while sulphuric acid consumption is simultaneously decreased by the addition of pyrite. Uranium concentrations in starting waste materials used for leaching were extremely low (0.0278 and 0.0372% U) but about 60% recovery of uranium was obtained, with relatively low consumption of sulphuric acid.