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The present study applied a non-invasive method to analyse anthropogenic particles and prey items in white stork (Ciconia ciconia) pellets. Pellets (n = 20) were obtained from white stork nests during the 2020 breeding season from two sites in Croatia. In total, 7869 anthropogenic particles were isolated. The majority of particles were fragments, while previous studies on other birds often reported fibres. An ATR–FTIR polymer analysis detected glass and construction and building materials, as well as several compounds associated with plastic masses. Polymer investigation revealed the presence of dotriacontane and octacosane, which are by-products of polyethylene (PE) degradation and transformation. Additionally, the detection of vinylidene chloride (VDC) highlights the historical contribution of polyvinylidene chloride (PVDC) to plastic pollution. Significant variation in particle quantity and size between the sampling sites was detected, with larger particles found at sites associated with the metal mechanical engineering industry and agriculture. Prey assessment revealed chitin remains of large insects such as Orthoptera and Coleoptera. This research confirms the potential of pellet analysis as a valuable tool for assessing the presence of anthropogenic particles in the environment. However, further research is needed to fully understand the extent of particle ingestion, particle sources and potential impact. [ABSTRACT FROM AUTHOR]

The present study assessed the effect of agricultural practices on biomarker response in white stork (Ciconia ciconia) nestlings from continental Croatia. During the breeding season of 2022, blood was sampled (n = 54) from the landfill Jakuševec and the agricultural areas Baranja and Srijem. Different patterns of biomarker response in regard to the sampling site were detected. Results demonstrate the presence of inhibitory pollutants from the landfill Jakuševec and agricultural area Baranja, which is reflected in lower cholinesterase (ChE) activity. Oxidative stress biomarkers show different responses concerning study sites. Glutathione reductase (GR) activity was higher in Baranja and Srijem, suggesting a different mixture of pollutants inducing an antioxidative response. Leachate from the landfill Jakuševec is suspected to affect the white stork nestlings by elevating the concentrations of reactive oxygen species (ROS). An environmental combination of pollutants appears to induce oxidative stress in white stork nestlings. In addition to agricultural practices, white stork nestlings may also be under environmental pressure from the surrounding pollution. Further research is warranted to include additional chemical analysis to associate the environmental concentrations with the potential adverse effects in apex predators, such as the white stork. [ABSTRACT FROM AUTHOR]

White stork nestlings can provide quantitative data on the quality of the environment, as they are dependent on their parents that provide locally foraged food. Blood was sampled from the brachial vein (n = 109) and the sampling was performed in parallel with ringing during breeding season 2020 from five areas in eastern Croatia: Lonjsko polje, Jelas polje, Slavonski Brod–east, Podunavlje, and Donje Podravlje. In the present study, for the first time in Croatia, the following enzymatic biomarkers were assessed in white stork nestlings: activities of acetylcholinesterase (AChE), carboxylesterase (CES), glutathione S‐transferase (GST), and glutathione reductase (GR), as well as nonenzymatic biomarkers: levels of glutathione (GSH) and reactive oxygen species (ROS). All endpoints were measured in two blood fractions: plasma and a postmitochondrial fraction (S9). Nestlings from Podunavlje and Donje Podravlje, areas known for intensive agriculture, showed lower AChE and CES activity when compared to the other investigated areas, indicating the presence of inhibitory xenobiotics. Higher oxidative stress was observed in Slavonski Brod–east, an area surrounded by metal and engineering industry, and Podunavlje compared to the other sampling areas. Hence, this study shows the impact of pollutants from the surrounding metal, petroleum, and agricultural industry might have on the biomarkers in white stork nestlings, which are often seen as early‐warning signals. Highlights: Plasma and S9 biomarkers were analyzed in white stork nestlings from Croatia. Lower acetylcholinesterase and carboxylesterase activities were recorded in agricultural areas. Higher oxidative stress (glutathione S‐transferase, glutathione reductase, glutathione, reactive oxygen species) is recorded in the metal industry area. Spatial variation in biomarker response may be related to local pollution. White stork nestlings appear to be affected by pollutants from their environment. [ABSTRACT FROM AUTHOR]

The goal of the present study was to investigate differences in biomarker responses related to metal(loid)s in white stork (Ciconia ciconia) nestling's blood from continental Croatia. To achieve this, a battery of biomarkers that can be affected by environmental pollutants, including metal(loid)s, was assessed (esterase activity, fluorescence–based oxidative stress biomarkers, metallothionein levels, glutathione–dependent enzyme activity). The research was conducted during the white stork breeding season in diverse areas (a landfill, industrial and agricultural sites, and an unpolluted area). White storks' nestlings near the landfill exhibited reduced carboxylesterase (CES) activity, elevated glutathione (GSH) concentration, as well as high Pb content in the blood. Increased As and Hg concentrations in blood were attributable to environmental contamination in agricultural area and an assumed unpolluted area, respectively. Furthermore, agricultural practices appeared to affect CES activity, as well as elevate Se levels. In addition to the successful implementation of biomarkers, present research showed that agricultural areas and a landfill are areas with increased metal(loid) levels possibly causing adverse effects on the white storks. This first–time heavy metal and metalloid analyses in the white stork nestlings from Croatia point to the necessary monitoring and future assessments of pollution impact to prevent irreversible adverse effects. [Display omitted] • White stork nestling blood (n = 106) was sampled in Pannonian Croatia. • Higher glutathione levels are associated with foraging at landfill Jakuševec. • Carboxylesterase activity is decreased in the agricultural area Donje Podravlje. • Nestlings had concerning levels of arsenic, selenium and lead in the blood. • Monitoring of white storks in Podunavlje, Crnac Polje and Jakuševec is necessary. [ABSTRACT FROM AUTHOR]

Simple summary: Protocols were adjusted and established for biomarker assessment in two blood fractions: plasma and the post-mitochondrial fraction (S9). Basal biomarker values were determined in white stork (Ciconia ciconia) nestlings from Croatia, and biomarker responses in two different types of blood samples were compared. Novel fluorescence-based glutathione and reactive oxygen species detection was established providing the potential usage of blood for assessment of environmental impact at the molecular level. White stork (Ciconia ciconia) nestlings can provide quantitative information on the quality of the surrounding environment by indicating the presence of pollutants, as they depend on locally foraged food. This study represents the first comparison of biomarkers in two fractions of white stork nestling blood: plasma and S9 (the post-mitochondrial fraction). The aim of this study was to evaluate acetylcholinesterase (AChE), carboxylesterase (CES), glutathione S-transferase (GST), and glutathione reductase (GR), as well as to establish a novel fluorescence-based method for glutathione (GSH) and reactive oxygen species (ROS) detection in plasma and S9. Considering the enzymatic biomarkers, lower variability in plasma was detected only for AChE, as CES, GST, and GR had lower variability in S9. Enzyme activity was higher in plasma for AChE, CES, and GST, while GR had higher activity in S9. Regarding the fluorescence-based method, lower variability was detected in plasma for GSH and ROS, although higher GSH detection was reported in S9, and higher ROS was detected in plasma. The present study indicated valuable differences by successfully establishing protocols for biomarker measurement in plasma and S9 based on variability, enzyme activity, and fluorescence. For a better understanding of the environmental effects on nestlings' physiological condition, biomarkers can be measured in plasma and S9. [ABSTRACT FROM AUTHOR]