Exposure to Phosphates and Nitrites through Meat Products: Estimation of the Potential Risk to Pregnant Women.

Diet during pregnancy is one of the most important nutritional challenges associated with some risks for the mother and the fetus. For the first time, the study aims to estimate long-term (2018-2022) exposure to nitrate and phosphates in Serbian pregnant women, based on individual consumption data and accurate values measured in frequently consumed meat products. For this purpose, seven types of meat products, consisting of 3047 and 1943 samples, were collected from retail markets across Serbia, to analyze nitrites and phosphorus content, respectively. These data were combined with meat product consumption data from the Serbian National Food Consumption Survey to assess dietary intake of nitrites and phosphate. The results were compared with the acceptable daily intake (ADI) proposed by the European Food Safety Authority. The average dietary exposure (EDI) to phosphorus ranged from 0.733 mg/kg bw/day (liver sausage and pate) to 2.441 mg/kg bw/day (finely minced cooked sausages). Considering nitrite intake, the major sources were bacon (0.030 mg/kg bw/day) and coarsely minced cooked sausages (0.0189 mg/kg bw/day). In our study, average nitrite and phosphorus exposure in the Serbian pregnant women population are far below the EFSA recommendations (ADI 0.07 mg/kg bw/day and 40 mg/kg bw/day, respectively).

Ochratoxin A potentiates citrinin accumulation in kidney and liver of rats.

Ochratoxin A (OTA) and citrinin (CTN) are nephrotoxic mycotoxins often found together in grain. The aim of this study was to measure their accumulation in the kidney and liver of adult male Wistar rats, see how it would be affected by combined treatment, and to determine if resveratrol (RSV) would decrease their levels in these organs. The rats received 125 or 250 mg/kg bw of OTA by gavage every day for 21 days and/or 20 mg/kg bw of CTN a day for two days. Two groups of rats treated with OTA+CTN were also receiving 20 mg/kg bw of RSV a day for 21 days. In animals receiving OTA alone, its accumulation in both organs was dose-dependent. OTA+CTN treatment resulted in lower OTA but higher CTN accumulation in both organs at both OTA doses. RSV treatment increased OTA levels in the kidney and liver and decreased CTN levels in the kidney. Our findings point to the competition between CTN and OTA for organic anion transporters 1 and 3.

The Intake of Phosphorus and Nitrites through Meat Products: A Health Risk Assessment of Children Aged 1 to 9 Years Old in Serbia.

This study provides the data on dietary exposure of Serbian children to nitrites and phosphorus from meat products by combining individual consumption data with available analytical data of meat products. A total of 2603 and 1900 commercially available meat products were categorized into seven groups and analysed for nitrite and phosphorous content. The highest mean levels of nitrite content, expressed as NaNO2, were found in finely minced cooked sausages (40.25 ± 20.37 mg/kg), followed by canned meat (34.95 ± 22.12 mg/kg) and coarsely minced cooked sausages (32.85 ± 23.25 mg/kg). The EDI (estimated daily intake) of nitrites from meat products, calculated from a National Food Consumption Survey in 576 children aged 1-9 years, indicated that the Serbian children population exceeded the nitrite ADI (acceptable daily intake) proposed by EFSA (European Food Safety Authority) in 6.4% of children, with a higher proportion in 1-3-year-old participants. The mean phosphorus concentration varied from 2.71 ± 1.05 g/kg to 6.12 ± 1.33 g/kg in liver sausage and pate and smoked meat products, respectively. The EDI of phosphorus from meat products was far below the ADI proposed by EFSA, indicating that the use of phosphorus additives in Serbian meat products is generally in line with legislation.

Dietary Exposure and Risk Assessment of Aflatoxin M1 for Children Aged 1 to 9 Years Old in Serbia.

The present study was conducted to estimate the exposure and characterize the risk for the child population of Serbia to Aflatoxin M1 (AFM1) from milk and milk-based food. A total of 3404 samples comprising milk and different milk-based food samples were collected from various regions of Serbia from 2017 to 2019. Evaluation of AFM1 exposure was carried out using the deterministic method, whereas risk characterization was evaluated using the margin of exposure (MOE) and the risk of hepatocellular carcinoma (HCC). Detection rates for AFM1 in milk and milk-based food samples ranged between 2% and 79%, with the highest incidence (79%) and mean level (22.34 ± 0.018 ng kg-1) of AFM1 being detected in pasteurized and UHT milk. According to the three consumption estimates, the values of estimated daily intake (EDI) were higher for toddlers as compared with children aged 3-9 years. Children aged 1-3 years had the highest risk of exposure to AFM1 in milk, with an estimated daily intake of 0.164 and 0.193 ng kg-1 bw day-1 using lower bound (LB) and upper bound (UB) exposure scenarios, respectively. Such difference could result from the higher consumption to weight in younger children. Based on the estimated daily intake (EDI) found in this study, the risk of AFM1 exposure due to consumption of milk and milk-based food was low since the MOE values obtained were >10,000. In addition, the risk of HCC cases/year/105 individuals of different age groups showed that the value of HCC, using potency estimates of 0.0017 (mean), was maximum (0.00034) in the age group 1-3 years, which indicates no health risk for the evaluated groups. The present study revealed the importance of controlling and preventing AFM1 contamination in milk through continuous monitoring and regular inspection to reduce the risk of AFM1 exposure, especially in children.

Efficacy of a Modified Clinoptilolite Based Adsorbent in Reducing Detrimental Effects of Ochratoxin A in Laying Hens.

BACKGROUND: The objective of this study was to evaluate the efficacy of modified clinoptilolite (Minazel Plus®, MZ) as a mycotoxin adsorbent for preventing the negative the effects of ochratoxin A (OTA) on performance, pathohistological changes, and OTA residue in the eggs of laying hens. METHODS: Forty eight (n = 48) laying hens (27 weeks old) were equally divided into six groups and depending on the type of addition were allocated to the following experimental treatments for 7 weeks: E-I group-1 mg/kg OTA; E-II group 0.25 mg/kg OTA; E-III group 1 mg/kg OTA + 0.2% of MZ; E-IV group 0.25 mg/kg OTA + 0.2% of MZ; MZ group supplemented with 0.2% of the adsorbent; and control (K, without feed additive). RESULTS: Overall, the addition of 0.2% MZ to laying hen feed mitigated the harmful effects of OTA on target organs and reduced the presence of OTA residue in eggs. The groups that received 0.2% of MZ achieved better production results in terms of body weight, number of eggs, and feed consumption, compared to the other treatments. CONCLUSIONS: The current findings confirm the efficacy of MZ in preventing performance losses in laying hens exposed to OTA, as well as for improving the welfare and health of food producing animals.

Presence of natural radionuclides and toxic elements in monocalcium phosphate, complete feed and pig manure.

Naturally occurring radionuclides and toxic elements are ubiquitous in the environment and can pose a great risk to animals' health. Control of these toxic substances in animal diet is essential for animal's welfare, human nutrition risk management as well as environmental protection. Our study addresses these issues by determining a content of natural radionuclides (40K, 238U and 226Ra), and toxic elements (Cd, Cr, As) in total of 108 samples: 36 monocalcium phosphate, 36 complete feed and 36 pig manure samples that were collected from 6 different commercial farms in the Republic of Serbia. In monocalcium phosphate the content of 40K, 238U and 226Ra ranged from 7.6-23.4 Bq kg-1, 13.2-1778 Bq kg-1 and 3.6-13.5 Bq kg-1, respectively. In complete feed samples, 40K ranged from 170 to 306 Bq kg-1, 238U from minimum detectable activity (MDA) to 9.1 Bq kg-1, and 226Ra from MDA-5.5 Bq kg-1, while in pig manure 40K ranged from 146 to 531 Bq kg-1, 238U from MDA-30.1 Bq kg-1, and 226Ra from MDA-10 Bq kg-1. The concentrations of Cd, Cr and As in monocalcium phosphate were 0.26-14.9 mg kg-1, 10.3-497 mg kg-1, and 4.2-5.5 mg kg-1, respectively. In complete feed, the content of Cd, Cr and As ranged from minimum detectable concentration (MDC) - 0.78 mg kg-1, 0.30-7.41 mg kg-1, and 0.09-0.31 mg kg-1, respectively. In pig manure Cd, Cr and As ranged from 0.8-2.3 mg kg-1, 2.3-22.2 mg kg-1, and 0.51-5.45 mg kg-1, respectively. The obtained results showed that Cd concentration in monocalcium phosphate exceeded the maximum permissible legislative concentrations for phosphate additives. Monocalcium phosphate is the main source of 238U and Cr in complete feed, while Cd derives from other sources. For that reason excessive use of monocalcium phosphate can lead to high levels of these pollutants in complete feed and consequently in manure.

Impact of climate change on aflatoxin M1 contamination of raw milk with special focus on climate conditions in Serbia.

BACKGROUND: The impact of climate change has been identified as an emerging issue worldwide owing to potential effects that can provoke changes in the nature and occurrence of food safety hazards. Mycotoxins are a group of naturally occurring toxic substances produced by several genera of filamentous fungi, among which, primarily aflatoxins, fumonisins, ochratoxins, trichothecenes and zearalenone are of the greatest concern owing to their negative impact on human health, animal productivity and international trade. RESULTS: Extreme weather conditions such as floods and droughts which have not occurred previously in Serbia may be supporting factors for contamination of crops by various species of toxigenic fungi and their related mycotoxins. In this study, principal component analysis (PCA) and multivariate analyses were used as powerful tools allowing better understanding of the range of climate factors impacting on crop contamination by mycotoxins. Among the selected environmental variables, temperature and moisture significantly influenced aflatoxin production. Recent drought and then flooding confirmed that Serbia is one of the few European countries with very high risk exposure to natural hazards, as well as that mycotoxins are one of the foodborne hazards most susceptible to climate change. Thus climate change effects on mycotoxigenic fungi and mycotoxin contamination are now receiving scientific attention, especially from a risk analysis perspective. CONCLUSION: This paper evaluates the available information on the influence of climate change on mycotoxin contamination, especially aflatoxin M1 (AFM1). In addition, this investigation should beneficially contribute to more accurate predictions of the regional risk from mycotoxins in future decades. © 2019 Society of Chemical Industry.

Effects of combined treatment with ochratoxin A and citrinin on oxidative damage in kidneys and liver of rats.

A multimycotoxin analysis approach in grains results in frequent simultaneous findings of nephrotoxic mycotoxins ochratoxin A (OTA) and citrinin (CTN). The mechanism of CTN and OTA toxicities in biological systems is not fully understood but it is known that oxidative stress is involved. In this study, oxidative damage of DNA, lipids, and the concentration of glutathione (GSH), as well as possible antioxidative effects of resveratrol (RSV) were studied in vivo. Male adult Wistar rats were treated orally with OTA (0.125 and 0.250 mg kg-1 b.w.), RSV (20 mg kg-1 b.w.) for 21 days, CTN (20 mg kg-1 b.w.) for two days or with their combinations. The hOGG1 modified comet assay revealed kidneys and liver oxidative DNA damage in OTA + CTN treated animals, which was not reversed by RSV. CTN did not reduce glutathione (GSH) or increase malondialdehyde (MDA) concentration in any tissue, while OTA reduced kidneys GSH and increased kidneys and liver MDA. RSV increased GSH concentrations in all tissues and decreased MDA concentration in the liver only. Oxidative stress is involved in the toxicity of OTA and CTN but it seems that it differs significantly in organs. Most of the effects on GSH and MDA in combined toxicity may be attributed to the toxic effects of OTA. RSV was effective in restoring the depleted GSH in all tissues but had no effect on the MDA concentration and DNA damage.

A review of the current situation of aflatoxin M1 in cow's milk in Serbia: risk assessment and regulatory aspects.

The aim of this systematic review is to provide information regarding the incidence and levels of aflatoxin M1 (AFM1) in raw and heat processed cow's milk in Serbia during 2015-16 and to compare these with collected data on the occurrence of AFM1 in raw milk and dairy products during the last decade in our region. Estimation of dietary exposure (EDI) and hazard index (HI) calculations for different age groups of the population were also carried out, based on the AFM1 content of milk samples and on available food consumption data in Serbia. AFM1 was detected in 69.9% (984/1408) of raw milk samples in 2015 versus 84.9% (3094/3646) in 2016, while in heat-processed milk, AFM1 was detected in 77.8% (364/468) in 2015 versus 98.5% (753/765) in 2016. On the basis of the obtained results, 450 (9%) of raw and 14 (1.1%) of heat-processed milk samples were contaminated with AFM1 levels above the maximum permitted level in Serbia (0.25 µg kg-1). However, a large percentage of raw and heat processed milk in Serbia (30.1% and 17.3%, respectively) was contaminated with AFM1 levels above the maximum permitted level regulated in the European Union (0.05 µg kg-1). Therefore, in order to protect consumer health, it is extremely important to further control the level of aflatoxins in milk, and this should be considered as a high priority for risk management actions.

A Blueprint to Evaluate One Health.

One Health (OH) positions health professionals as agents for change and provides a platform to manage determinants of health that are often not comprehensively captured in medicine or public health alone. However, due to the organization of societies and disciplines, and the sectoral allocation of resources, the development of transdisciplinary approaches requires effort and perseverance. Therefore, there is a need to provide evidence on the added value of OH for governments, researchers, funding bodies, and stakeholders. This paper outlines a conceptual framework of what OH approaches can encompass and the added values they can provide. The framework was developed during a workshop conducted by the "Network for Evaluation of One Health," an Action funded by the European Cooperation in Science and Technology. By systematically describing the various aspects of OH, we provide the basis for measuring and monitoring the integration of disciplines, sectors, and stakeholders in health initiatives. The framework identifies the social, economic, and environmental drivers leading to integrated approaches to health and illustrates how these evoke characteristic OH operations, i.e., thinking, planning, and working, and require supporting infrastructures to allow learning, sharing, and systemic organization. It also describes the OH outcomes (i.e., sustainability, health and welfare, interspecies equity and stewardship, effectiveness, and efficiency), which are not possible to obtain through sectoral approaches alone, and their alignment with aspects of sustainable development based on society, environment, and economy.

Comparative investigation of the efficacy of three different adsorbents against OTA-induced toxicity in broiler chickens.

The aim of our study was to determine the efficacy of three different adsorbents, inorganic (modified zeolite), organic (esterified glucomannans) and mixed (inorganic and organic components, with the addition of enzymes), in protecting broilers from the toxic effects of ochratoxin A in feed. Broilers were fed diets containing 2 mg/kg of ochratoxin A (OTA) and supplemented with adsorbents at the recommended concentration of 2 g/kg for 21 days. The presence of OTA led to a notable reduction in body weight, lower weight gain, increased feed conversion and induced histopathological changes in the liver and kidneys. The presence of inorganic, organic and mixed adsorbents in contaminated feed only partially reduced the negative effects of OTA on the broiler performances. Broilers that were fed with adsorbent-supplemented feed reached higher body weight (17.96%, 19.09% and 13.59%), compared to the group that received only OTA. The presence of adsorbents partially alleviated the reduction in feed consumption (22.68%, 12.91% and 10.59%), and a similar effect was observed with feed conversion. The applied adsorbents have also reduced the intensity of histopathological changes caused by OTA; however, they were not able to prevent their onset. After the withdrawal of the toxin and adsorbents from the feed (21-42 days), all previously observed disturbances in broilers were reduced, but more remarkably in broilers fed with adsorbents.

Cadmium and mercury accumulation in European hare (Lepus europaeus): age-dependent relationships in renal and hepatic tissue.

A total of 63 European hares have been collected from five Serbian agricultural regions. The hares assayed were divided into four age groups (3-6 months, 12 months, 12-24 months, and 24-36 months) and investigated upon presence at cadmium (Cd) and mercury (Hg) in the kidney and liver. The positive significant correlation (Ps-Pearson's coefficient) between Cd concentrations in the kidney and liver within age group the 3-6 months was found (Ps = 0.81, p < 0.001). Differences between Cd content in the kidney in comparison to liver were significant within all presented age groups (p < 0.001). Differences between recorded Hg levels in the kidney were not significant between presented age groups (p > 0.05). Statistically significant differences were registered between Hg content in the liver of the hares aged 24-36 and 12 months (p < 0.05). There were no statistically significant correlations registered between Hg concentrations in the kidney and liver within any particular age group (p > 0.05). The strong statistically significant associations were registered between Cd and Hg content in the liver (Cd L/Hg L) in the age group 3-6 and 12-24 months (Cd L/Hg L, Ps = 0.94; p < 0.001 and Ps = 0.91; p < 0.001, respectively). The polynomial regression model used for graphing the observed data seems to be a method for modeling the relationship between measured Cd and Hg concentrations in the liver and kidney as first approximation for bioaccumulation in hares.

The role of total fats, saturated/unsaturated fatty acids and cholesterol content in chicken meat as cardiovascular risk factors.

BACKGROUND: The objective of the study was to present information about the chemical composition, the fatty acids profile, and cholesterol content of chicken meat in order to investigate the impact of chicken meat consumption on cardiovascular risk in the general population. METHODS: A total of 48 6-wk-old broiler chickens broilers from two farms in June to November of 2012, and February of 2013, were used in this trial. Total lipid content was determined by extraction of fat by petrol ether (Soxhlet) after acid hydrolysis of samples. Fatty acids were determined by capillary gas chromatography. Cholesterol determination was performed by using HPLC/PDA system. RESULTS: The results indicate that the total free cholesterol content in raw breast and drumstick of chickens was in the range of 37,41-79,9 mg/100 g and 48,35-99,5 mg/100 g, respectively. The main fatty acids identified in all cuts were C18:1c9, C18:2n6, C16:0, C18:0, and C16:1. Decreasing the dietary n-6/n-3 clearly decreased the content in breast and drumstick muscle of C18:2n6, C18:3n3, and C20: 3n6, but increased that of C16:0, C18:0, and C20:2. Also, the major saturated fatty acid (SFA) (C16:0 and C18:0) was significantly differ among the four treatments. CONCLUSION: Our study shows that dietary fat and fatty acid composition influence the concentrations of total cholesterol content, total fat content, and fatty acid composition in broiler muscle. This information will aid in determining the burden of chicken meat as a cardiovascular risk factors disease and act as a planning tool for public-health Programmes.

Agent-based assessment of stormwater re-use potential of low-impact development control facilities at the site of Vlasina Lake, Serbia.

Vlasina Lake in south-east Serbia is classified as an Area of Distinct Land Use and, as such, is subject to high environmental protection standards applied in the Master Plan. Two open channels for stormwater and sediment transportation to two large detention basins with pumping stations for water evacuation into the lake were envisaged in the Master Plan. In the preliminary design, the stormwater system was quite different: wherever possible, on-site natural features were used for allocation of ponds, and drainage channels were led through existing road culverts. The applied design concept has been low impact development (LID), which led to potential blue-green corridors, recognized by project stakeholders. The paper studies the possibility of using ponds as a key element of both the LID concept and the blue-green corridors approach. For that purpose, an initial Vlasina Lake site agent-based simulation model has been created. A realistic physical model is included, and simulation results for two hypothetical climatic and socio-economic scenarios are presented. From the experience in creating the agent-based model, and based on the simulation results, recommendations are given for further work. It is shown that ponds have potential for the investigated water re-use purposes.

Environmental Cd and Zn concentrations in liver and kidney of European hare from different Serbian regions: age and tissue differences.

A total of 84 European hares collected from eleven Serbian regions investigated upon cadmium (Cd) and zinc (Zn) presence. Strong statistically significant correlations between Cd concentrations in kidney and liver were registered in animals older than 12 months. Significant differences between Zn concentrations in liver in comparison to kidney were found within every single age group with exception of the oldest. Negative correlation (Ps-Pearson's correlation) between Zn and Cd concentrations were found in liver samples within the age group of 12 months (Ps = -0.67, p = 0.004).

A survey of spontaneous occurrence of ochratoxin A residues in chicken tissues and concurrence with histopathological changes in liver and kidneys.

Toxicological and histopathological investigations of tissues of commercially slaughtered chickens were carried out to provide a preliminary evaluation of the incidence of occurrence of ochratoxin A (OTA) in chicken sold in Serbian retail market. In addition, the etiology of nephropathies of these chickens was elucidated. The majority of these tissue samples were not found to contain measurable amounts of OTA. Moreover, the OTA levels found in analyzed tissues were generally low and there was no positive correlation between the presence of OTA and the frequency of histopathological changes. Histopathological changes such as degenerative changes in the kidneys and liver differed from the classical description of the mycotoxic nephropathy, indicating that the chicken nephropathy observed in Serbia may have a multitoxic etiology with possible synergistic effect between microorganisms and natural toxins, usually present in low concentrations. The low OTA results also suggested that chicken meat available in the retail market in Serbia are unlikely to pose any significant adverse health risk to the consumers with respect to OTA toxicity.

WITHDRAWN: Comparison of high performance liquid chromatography and liquid chromatography/tandem mass spectrometry methods in the quantification and confirmation of ochratoxin A in pig tissue.

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Real and perceived risks for mycotoxin contamination in foods and feeds: challenges for food safety control.

Mycotoxins are toxic compounds, produced by the secondary metabolism of toxigenic moulds in the Aspergillus, Alternaria, Claviceps, Fusarium, Penicillium and Stachybotrys genera occurring in food and feed commodities both pre- and post-harvest. Adverse human health effects from the consumption of mycotoxins have occurred for many centuries. When ingested, mycotoxins may cause a mycotoxicosis which can result in an acute or chronic disease episode. Chronic conditions have a much greater impact, numerically, on human health in general, and induce diverse and powerful toxic effects in test systems: some are carcinogenic, mutagenic, teratogenic, estrogenic, hemorrhagic, immunotoxic, nephrotoxic, hepatotoxic, dermotoxic and neurotoxic. Although mycotoxin contamination of agricultural products still occurs in the developed world, the application of modern agricultural practices and the presence of a legislatively regulated food processing and marketing system have greatly reduced mycotoxin exposure in these populations. However, in developing countries, where climatic and crop storage conditions are frequently conducive to fungal growth and mycotoxin production, much of the population relies on subsistence farming or on unregulated local markets. Therefore both producers and governmental control authorities are directing their efforts toward the implementation of a correct and reliable evaluation of the real status of contamination of a lot of food commodity and, consequently, of the impact of mycotoxins on human and animal health.

Evaluation and validation of two chromatographic methods (HPLC-fluorescence and LC-MS/MS) for the determination and confirmation of ochratoxin A in pig tissues.

Two different analytical methods for the determination and confirmation of ochratoxin A (OTA) in blood serum, kidney, and liver of pigs have been compared. Sample cleanup was based on liquid-liquid phase extraction. The detection of OTA was accomplished with high-performance liquid chromatography (HPLC) combined either with fluorescence detection (FLD) or electrospray ionization (ESI+) tandem mass spectrometry (MS/MS). The comparative method of evaluation was based on the investigation of 90 samples of blood serum, kidney, and liver per animal originating from different regions of Serbia. The analytical results are discussed in view of the respective method validation data and the corresponding experimental protocols. In general, analytical data obtained with liquid chromatography (LC)-MS/MS detection offered comparable good results in the sub-ppb concentration level indicating that the electrospray tandem mass spectrometric (LC-MS/MS) method was more selective and sensitive for the analysis and confirmation of OTA in pig tissues than the HPLC method after the methylation of OTA.

Evaluation and validation of two different chromatographic methods (HPLC and LC-MS/MS) for the determination and confirmation of ochratoxin A in pig tissues.

This study was undertaken to compare two different analytical methods for the determination and confirmation of ochratoxin A (OTA) in blood serum, kidney and liver of pigs. Sample clean-up was based on liquid-liquid phase extraction. The detection of OTA was accomplished with high-performance liquid chromatography (HPLC) combined either with fluorescence detection (FL) or electro spray ionization (ESI+) tandem mass spectrometry (MS-MS). Comparative method evaluation was based on the investigation of 90 samples of blood serum, kidney and liver per animal originating from different regions of Serbia. The analytical results are discussed in view of the respective method validation data and the corresponding experimental protocols. In general, analytical data obtained with (LC-MS-MS) liquid chromatography electro spray tandem mass spectro metry detection offered comparable good results at the sub-ppb concentration level. The results indicate that the liquid chromatography electro spray tandem mass spectrometric (LC-MS/MS) method was more specific and sensitive for the analysis and confirmation of ochratoxin A in pig tissues then high pressure liquid chromatography (HPLC) method after methylation of OTA.