Microbiological Assessment of Dairy Products Produced by Small-Scale Dairy Producers in Serbia.

The microbiological quality of dairy products from small-scale producers in Serbia was analysed. A total of 302 dairy products [raw (n = 111) and pasteurized milk cheeses (n = 79) and kajmak (n = 112)], were collected and tested for the presence of pathogens, Listeria monocytogenes and Salmonella spp., and enumerated for Coagulase-positive staphylococci (CPS), Escherichia coli, and yeasts and moulds. None of the samples tested positive for Salmonella spp., while L. monocytogenes was recovered from one raw milk cheese and five kajmak samples. Raw milk cheese and kajmak also had higher levels of indicator microorganisms, namely E. coli and yeast and moulds. Molecular serotyping grouped L. monocytogenes isolates into serogroups 1 (1/2a and 3a) and 3 (1/2b, 3b, and 7). When exposed to eight antibiotics, L. monocytogenes isolates were mostly sensitive, with the exception of oxacillin and reduced susceptibility to clindamycin, penicillin G, and trimethoprim/sulfamethoxazole, emphasizing the importance of continuous surveillance for antimicrobial resistance. Samples that tested positive for Listeria spp. also had higher loads of indicator microorganisms, namely E. coli and yeast and moulds, suggesting lapses in hygiene practices during production. Collectively, these data emphasize the need for improved food safety and hygiene practices among small-scale dairy producers. This is crucial to reduce the microbial contamination and improve both the quality and safety of dairy products in the Serbian market.

Cumulative risk assessment of dietary exposure of the adult population in Serbia to pesticides that have chronic effects on the thyroid gland through fresh fruits and vegetables.

In contrast to the traditional approach to risk assessment, which focuses on a single chemical, cumulative exposure and risk assessment considers the consequences of exposure to multiple chemical combinations. A cumulative risk assessment of dietary exposure of adult females and adult males to pesticides with chronic effects on the thyroid was conducted by estimation of the Total Margin of Exposure (MOET). Exposure to each active substance was estimated using a second-order Monte Carlo simulation. Input values for the simulation were based on over 2300 conventionally produced fruit and vegetable samples analysed from 2021 to 2023 and consumption data collected using the Food Frequency Questionnaire (FFQ) and 24h recall method. MOET values, for both populations assessed, did not exceed thresholds for regulatory consideration established by risk managers. Considering that MOETs values from consumption of fresh fruits and vegetables were relatively close to the threshold value, total exposure to the pesticides could likely reach risk-associated MOET levels, through the consumption of other food products that may be contaminated.

Second order probabilistic assessment of chronic dietary exposure to aflatoxin M1 in Serbia.

Considering the genotoxic and cancerogenic nature of aflatoxin M1 (AFM1), its presence in milk and dairy products may pose health risks for consumers. The chronic exposure was calculated using a two-dimensional (second order) Monte Carlo model. Results of 13 722 milk and dairy product samples analysed in the 2015-2022 period were used. Milk and dairy products intake information was collected with a Food Frequency Questionnaire (FFQ) validated by a 24-h recall-based method. Risk characterization was done by calculation of the Margin of Exposure (MOE) and by calculation of AFM1 induced number of hepatocellular carcinoma (HCC) cases. Mean AFM1 Estimated Daily Intake (EDI) was highest in children at 0.336 (CI: 0.294-0.385) ng kg-1 bw day-1, followed by adolescents with 0.183 (CI: 0.164-0.204), then adult females with 0.161 (CI: 0.146-0.179) and finally adult males with lowest EDI of 0.126 (CI: 0.115-0.139) ng kg-1 bw day-1. MOE values based on mean EDI for all population groups were above risk associated threshold and the number of possible HCC cases was in the range of 0.0002-0.0021 cases per year for 105 individuals. The results suggest low health risks due to AFM1 exposure for the whole population. Still, this risk is not non-existent, especially for children as they have a higher ratio of the population exposed to risk associated AFM1 levels, with MOE values below risk indicating threshold starting at 77.5th percentile.

Quality and Oral Processing Characteristics of Traditional Serbian Cevap Influenced by Game Meat.

This study analyzes the influence of two different types of game meat (deer and wild boar) in relation to quality characteristics and oral processing attributes of skinless sausage. The goal of this study was to compare grilled game-meat-based "cevap" with conventional pork-meat-based samples. Research comprised of color analysis, evaluation of textural components, testing degree of difference, temporal dominance of sensations, calculation of main oral processing attributes, and examination of particle size distribution. The results show that oral processing attributes are similar in between samples and concur with results of the pork-based sample. This confirms the working hypothesis that it is possible to make game-meat-based "cevap" fully comparable with conventional pork meat products. In parallel, color and flavor characteristics are influenced by the type of game meat in the sample. Most of the dominant sensory attributes that occurred during mastication were game meat flavor and juiciness.

Sandwich ELISA for the Quantification of Nucleocapsid Protein of SARS-CoV-2 Based on Polyclonal Antibodies from Two Different Species.

In this study, a cost-effective sandwich ELISA test, based on polyclonal antibodies, for routine quantification SARS-CoV-2 nucleocapsid (N) protein was developed. The recombinant N protein was produced and used for the production of mice and rabbit antisera. Polyclonal N protein-specific antibodies served as capture and detection antibodies. The prototype ELISA has LOD 0.93 ng/mL and LOQ 5.3 ng/mL, with a linear range of 1.52-48.83 ng/mL. N protein heat pretreatment (56 °C, 1 h) decreased, while pretreatment with 1% Triton X-100 increased analytical ELISA sensitivity. The diagnostic specificity of ELISA was 100% (95% CI, 91.19-100.00%) and sensitivity was 52.94% (95% CI, 35.13-70.22%) compared to rtRT-PCR (Ct < 40). Profoundly higher sensitivity was obtained using patient samples mostly containing Wuhan-similar variants (Wuhan, alpha, and delta), 62.50% (95% CI, 40.59 to 81.20%), in comparison to samples mostly containing Wuhan-distant variants (Omicron) 30.00% (6.67-65.25%). The developed product has relatively high diagnostic sensitivity in relation to its analytical sensitivity due to the usage of polyclonal antibodies from two species, providing a wide repertoire of antibodies against multiple N protein epitopes. Moreover, the fast, simple, and inexpensive production of polyclonal antibodies, as the most expensive assay components, would result in affordable antigen tests.

Risk assessment of dietary exposure to aflatoxin B1 in Serbia.

Classified as Group 1 carcinogens aflatoxins (AFs) are of major concern for food safety and public health. Within the AFs group, aflatoxin B1 (AFB1) occurs at the highest levels in different food products and is considered as the most potent representative. A total of 463 samples of products susceptible to contamination with AFB1 and commonly consumed in Serbia were collected and analyzed. Consumption surveys were performed using a food frequency questionnaire (FFQ) and 24-h recall-based method. Monte Carlo simulation of 100,000 iterations was performed to estimate the intake of AFB1 through each food category, while a simple distribution approach was performed to estimate overall exposure of tested population groups. Risk characterization was performed by calculation of the Margin of Exposure (MOE) and by calculation of the number of possible hepatocellular carcinoma (HCC) cases. The highest exposure was observed in children with the mean overall estimated daily intake (EDI) in the range of 0.79-1.10 and 1.20-1.66 ng kg-1 bw day-1, followed by adolescents 0.56-0.81 and 0.94-1.32 ng kg-1 bw day-1, adult females 0.52-0.72 and 0.56-0.76 ng kg-1 bw day-1 and adult males with the mean overall EDI in the range of 0.39-0.56 and 0.47-0.66 ng kg-1 bw day-1 depending on consumption survey method and for the lower (LB) and upper bound (UB) scenario. MOE values that resulted from the mean values of the overall EDI of AFB1 were low, indicating high concern for all population groups. Based on obtained mean values of the overall EDI higher estimates of possible HCC cases caused by exposure to AFB1 were in the range of 0.01-0.02 cases/year/105 individuals, depending on the population group.

Comparison of Supercritical CO2-Drying, Freeze-Drying and Frying on Sensory Properties of Beetroot.

The aim of this study was to compare the sensory quality and acceptance of dried ready-to-eat beetroot snacks as a result of different drying methods applied: supercritical CO2-drying (scCO2-drying), frying, and freeze-drying. Descriptive sensory analysis, quality rating (10 assessors), and consumer acceptance testing (n = 102) were performed. Mean overall quality scores within the range of "very good" quality were found only in non-precooked scCO2-dried samples which were characterized by typical magenta color, low level of shape and surface deformations, pronounced brittleness and crispiness, and good rehydration during mastication. The other samples were in the range of "good" quality. The pre-cooking step before scCO2-drying negatively influenced the sensory quality parameters, particularly appearance. Around 60% of tested consumers showed a preference for the fried and non-precooked scCO2-dried samples. The drivers of liking were mostly related to the characteristics of the product, which was salted, fried, and crispy, with an oily and overburnt flavor, i.e., the product most similar to commercial potato chips products. Freeze-drying had a negative effect primarily on appearance and flavor. According to the sensory evaluation conducted, direct scCO2-drying without a pre-cooking step showed itself as a promising alternative drying technology in the production of dried beetroot snacks.

Water safety plan enhancements with improved drinking water quality detection techniques.

Drinking water quality has been regulated in most European countries for nearly two decades by the drinking water directive 98/83/EC. The directive is now under revision with the goal of meeting stricter demands for safe water for all citizens, as safe water has been recognized as a human right by the United Nations. An important change to the directive is the implementation of a risk-based approach in all regulated water supplies. The European Union Framework Seventh Programme Aquavalens project has developed several new detection technologies for pathogens and indicators and tested them in water supplies in seven European countries. One of the tasks of the project was to evaluate the impact of these new techniques on water safety and on water safety management. Data were collected on risk factors to water safety for five large supplies in Denmark, Germany, Spain and the UK, and for fifteen small water supplies in Scotland, Portugal and Serbia, via a questionnaire aiming to ascertain risk factors and the stage of implementation of Water Safety Plans, and via site-specific surveys known as Sanitary Site Inspection. Samples were collected from the water supplies from all stages of water production to delivery. Pathogens were detected in around 23% of the 470 samples tested. Fecal contamination was high in raw water and even in treated water at the small supplies. Old infrastructure was considered a challenge at all the water supplies. The results showed that some of the technique, if implemented as part of the water safety management, can detect rapidly the most common waterborne pathogens and fecal pollution indicators and therefore have a great early warning potential; can improve water safety for the consumer; can validate whether mitigation methods are working as intended; and can confirm the quality of the water at source and at the tap.

Exposure Assessment and Risk Characterization of Aflatoxin M1 Intake through Consumption of Milk and Yoghurt by Student Population in Serbia and Greece.

The objective of this research was to perform an exposure assessment of aflatoxin M1 (AFM1) intake through the consumption of milk and yoghurt by the student population in Serbia and Greece. A food consumption survey of milk and yoghurt was performed during the first half of 2018 in the two countries with at least 500 interviewees (aged between 18 and 27 years) per country, covering their dietary habits and body weight based on one-day and seven-day recall methods. Values for the concentration of AFM1 were extracted from published research. Finally, a Monte Carlo analysis of 100,000 iterations was performed to estimate the intake of AFM1 from the consumption of the two dairy products. Results revealed that the estimated average exposure of students to AFM1 was in the range of 1.238⁻2.674 ng kg-1 bw day-1 for Serbia, and 0.350⁻0.499 ng kg-1 bw day-1 for Greece, depending on the dietary recall method employed. High estimations for hepatocellular carcinoma (HCC) cases/year/105 individuals, depending on the prevalence of Hepatitis B virus surface antigen positive individuals (HBsAg+), were 0.0036⁻0.0047 and 0.0007⁻0.0009 for Serbia and Greece, respectively. Presented Margin of Exposure (MOE) and Hazard Index (HI) values indicate increased risk from exposure to AFM1, particularly in Serbia.

Exposure assessment and risk characterization of aflatoxins intake through consumption of maize products in the adult populations of Serbia, Croatia and Greece.

The main objective of this research was to perform an exposure assessment of aflatoxins intake through consumption of maize products in Serbia, Croatia, and Greece. Food consumption survey of maize-based products has been performed during 2017 in the three countries with at least 1,000 interviewees per country covering their dietary habits and body weight. Values for the concentration of aflatoxins were extracted from available research published in the last ten years. Finally, a Monte Carlo analysis of 100,000 iterations was performed to estimate the intake of aflatoxins from consumption of maize-based products. Results revealed that the estimated average exposure of adults to aflatoxins, from maize consumption, in each of the three countries was between 0.44 ng kg-1 bw day-1 and 5.59 ng kg-1 bw day-1. Margin of exposure values for the mean exposure levels, in all three countries, were between 30 and 389. Estimations for hepatocellular carcinoma cases/year/105 individuals, depending on the HBsAg+ prevalence, were 0.075-0.098, 0.006-0.008 and 0.020-0.026 for Serbia, Croatia and Greece, respectively.

Overview on the Mycotoxins Incidence in Serbia in the Period 2004⁻2016.

With an average annual production of 6.9 M tonnes and 2.5 M tonnes of maize and wheat respectively, Serbia is one of the main grain producers and exporters in Europe. Cereals are also the major staple food in Serbian diet. In view of the high cereal consumption, for human and animal nutrition, the presence of mycotoxins entails a high public health risk of chronic exposure to mycotoxins. This study provides an overview of the incidence of predominant mycotoxins, mainly in cereal and dairy products, in Serbia, in the 2004⁻2016, using data reported in the scientific literature. The study demonstrated that the total prevalence of aflatoxins was 62.9% (n = 12,517) with 26.2% of the samples exceeding the EU limits during this period. Results obtained for T-2/HT-2 (n = 523), deoxynivalenol (n = 2907), fumonisins (n = 998), zearalenone (n = 689) and ochratoxin A (n = 740) indicated the prevalence of 45.5%, 42.9%, 63.3%, 39.3% and 28.1%, respectively. For these mycotoxins, the EU limits were less frequently exceeded. Comprehensive collection and analysis of all accessible information reviewed in this paper showed moderate incidence and prevalence of mycotoxins in Serbia, with an exception of the 2012 drought year and the 2014 flood year.

Oxygen Consumption Rate Analysis of Mitochondrial Dysfunction Caused by Bacillus cereus Cereulide in Caco-2 and HepG2 Cells.

The emetic syndrome of Bacillus cereus is a food intoxication caused by cereulide (CER) and manifested by emesis, nausea and in most severe cases with liver failure. While acute effects have been studied in the aftermath of food intoxication, an exposure to low doses of cereulide might cause unnoticed damages to the intestines and liver. The toxicity which relies on the mitochondrial dysfunction was assessed on Caco-2 and HepG2 cells after exposure of one, three and ten days to a range of low doses of cereulide. Oxygen consumption rate analyses were used to study the impact of low doses of CER on the bioenergetics functions of undifferentiated Caco-2 and HepG2 cells using Seahorse XF extracellular flux analyzer. Both Caco-2 and HepG2 cells experienced measurable mitochondrial impairment after prolonged exposure of 10 days to 0.25 nM of cereulide. Observed mitochondrial dysfunction was greatly reflected in reduction of maximal cell respiration. At 0.50 nM CER, mitochondrial respiration was almost completely shut down, especially in HepG2 cells. These results corresponded with a severe reduction in the amount of cells and an altered morphology, observed by microscopic examination of the cells. Accurate and robust quantification of basal respiration, ATP production, proton leak, maximal respiration, spare respiratory capacity, and non-mitochondrial respiration allowed better understanding of the effects of cereulide in underlying respiratory malfunctions in low-dose exposure.