The presymptomatic treatment with 3HFWC nanosubstance decreased plaque load in 5XFAD mouse model of Alzheimer's disease.

INTRODUCTION: In the present study, we assessed the effects of the hyper-harmonized-hydroxylated fullerene-water complex (3HFWC) on Alzheimer's disease (AD) neuropathological hallmarks in 5XFAD mice, an AD animal model. METHODS: The 3-week-old 5XFAD mice were exposed to 3HFWC water solution ad libitum for 3 months in the presymptomatic phase of pathology. The functional effects of the treatment were confirmed through near-infrared spectroscopy (NIRS) analysis through machine learning (ML) using artificial neural networks (ANNs) to classify the control and 3HFWC-treated brain tissue samples. The effects of 3HFWC treatment on amyloid-ß (Aß) accumulation, plaque formation, gliosis, and synaptic plasticity in cortical and hippocampal tissue were assessed. RESULTS: The 3HFWC treatment significantly decreased the amyloid-ß plaque load in specific parts of the cerebral cortex. At the same time, 3HFWC treatment did not induce the activation of glia (astrocytes and microglia) nor did it negatively affect synaptic protein markers (GAP-43, synaptophysin, and PSD-95). CONCLUSION: The obtained results point to the potential of 3HFWC, when applied in the presymptomatic phase of AD, to interfere with amyloid plaque formation without inducing AD-related pathological processes such as neuroinflammation, gliosis, and synaptic vulnerability.

The Expression of Major Facilitator Superfamily Domain-Containing Protein2a (Mfsd2a) and Aquaporin 4 Is Altered in the Retinas of a 5xFAD Mouse Model of Alzheimer's Disease.

Cerebral amyloid angiopathy (CAA) is characterized by amyloid ß (Aß) accumulation in the blood vessels and is associated with cognitive impairment in Alzheimer's disease (AD). The increased accumulation of Aß is also present in the retinal blood vessels and a significant correlation between retinal and brain amyloid deposition was demonstrated in living patients and animal AD models. The Aß accumulation in the retinal blood vessels can be the result of impaired transcytosis and/or the dysfunctional ocular glymphatic system in AD and during aging. We analyzed the changes in the mRNA and protein expression of major facilitator superfamily domain-containing protein2a (Mfsd2a), the major regulator of transcytosis, and of Aquaporin4 (Aqp4), the key player implicated in the functioning of the glymphatic system, in the retinas of 4- and 12-month-old WT and 5xFAD female mice. A strong decrease in the Mfsd2a mRNA and protein expression was observed in the 4 M and 12 M 5xFAD and 12 M WT retinas. The increase in the expression of srebp1-c could be at least partially responsible for the Mfsd2a decrease in the 4 M 5xFAD retinas. The decrease in the pericyte (CD13+) coverage of retinal blood vessels in the 4 M and 12 M 5xFAD retinas and in the 12 M WT retinas suggests that pericyte loss could be associated with the Mfsd2a downregulation in these experimental groups. The observed increase in Aqp4 expression in 4 M and 12 M 5xFAD and 12 M WT retinas accompanied by the decreased perivascular Aqp4 expression is indicative of the impaired glymphatic system. The findings in this study reveal the impaired Mfsd2a and Aqp4 expression and Aqp4 perivascular mislocalization in retinal blood vessels during physiological (WT) and pathological (5xFAD) aging, indicating their importance as putative targets for the development of new treatments that can improve the regulation of transcytosis or the function of the glymphatic system.

American Foulbrood-Old and Always New Challenge.

American foulbrood (AFB) is exclusively an infectious disease of honey bee larvae (Apis mellifera) and their subspecies that is spread easily and rapidly and is often present in apiaries. Due to the resistance and pathogenicity of the bacterial causative agent of the disease, which has considerable epizootiological and economic significance for beekeeping, AFB was classified as a highly dangerous, infectious animal disease by the World Organization for Animal Health (WOAH). Considering the severity of the infection, a frequent occurrence, rapid and easy spread, epizooty and enzooty are common. We tried to present an overview of the latest information related to AFB through several chapters. In addition to the latest data on the etiology of the causative agent, the most important elements of the clinical signs of the disease are also listed. Along with an overview of classic microbiological and the latest molecular methods of diagnosis, we also discuss AFB treatment from its differential diagnostic aspect. We hope that through demonstrating the mentioned preventive measures and measures of good beekeeping practice, the review will contribute to the preservation of the health of bees and therefore the overall biodiversity of the planet.

Food Restriction Counteracts Dexamethasone-Induced Downregulation of Genes Involved in Cholesterol Homeostasis in Rat Brain during Aging.

Glucocorticoids are the most potent anti-inflammatory agents known. Limited in vivo data are available to characterize the mechanism underlying their cognitive side effects and transient occurrence of steroid psychosis. Cholesterol is important for proper neurotransmission and brain plasticity, and disruption of its homeostasis in the brain has been closely associated with memory decline during aging and in age-related neurodegenerative disorders. In the present study, we assessed the direct effects of dexamethasone, a potent synthetic glucocorticoid, on the expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), apolipoprotein E (ApoE) and cholesterol 24S-hydroxylase (CYP46A1), major enzymes involved in cholesterol synthesis, metabolism, and excretion, respectively. The effects of the dexamethasone were examined during aging, in the cortex and hippocampus of 6-, 12- and 18-month-old rats, and following long-term food restriction (FR). The most prominent change observed was the age-related decrease in ApoE mRNA regardless of the food regimen applied. In animals kept on FR, this decrease was accompanied by an increase in the mRNA expression of HMGCR and CYP46A1. The present study also demonstrates that food restriction reversed most of the dexamethasone-induced changes in the expression of genes involved in regulation of cholesterol homeostasis in aging rats, in a region-specific manner.

Corticosterone and Glucocorticoid Receptor in the Cortex of Rats during Aging-The Effects of Long-Term Food Restriction.

Numerous beneficial effects of food restriction on aging and age-related pathologies are well documented. It is also well-established that both short- and long-term food restriction regimens induce elevated circulating levels of glucocorticoids, stress-induced hormones produced by adrenal glands that can also exert deleterious effects on the brain. In the present study, we examined the effect of long-term food restriction on the glucocorticoid hormone/glucocorticoid receptor (GR) system in the cortex during aging, in 18- and 24-month-old rats. Corticosterone level was increased in the cortex of aged ad libitum-fed rats. Food restriction induced its further increase, accompanied with an increase in the level of 11ß-hydroxysteroid dehydrogenase type 1. However, alterations in the level of GR phosphorylated at Ser232 were not detected in animals on food restriction, in line with unaltered CDK5 level, the decrease of Hsp90, and an increase in a negative regulator of GR function, FKBP51. Moreover, our data revealed that reduced food intake prevented age-related increase in the levels of NFκB, gfap, and bax, confirming its anti-inflammatory and anti-apoptotic effects. Along with an increase in the levels of c-fos, our study provides additional evidences that food restriction affects cortical responsiveness to glucocorticoids during aging.

Honey Bees and Their Products as Indicators of Environmental Element Deposition.

The aim of this study was to evaluate element (sodium, magnesium, potassium, calcium, chromium, manganese, iron, cobalt, nickel, copper, zinc, arsenic, cadmium, and lead) deposition in honey bees (Apis mellifera L.) (worker bees, drone bees, and bee broods) and their products (wax and multifloral honey) in the central and north parts of Serbia using inductively coupled plasma mass spectrometry (ICP-MS). The study was carried out during the spring and summer seasons when the honeybees were active (2019). Fifty-four colonies of honey bees from different apiaries (located in Rudnik, Lazarevac, and Lezimir) were used in this study. Significant differences in element concentrations were found among locations (P < 0.05). The highest deposition of elements (sodium, calcium, manganese, iron, cobalt, nickel, zinc, arsenic, and cadmium) was found in worker bees. The most commonly detected elements in wax were chromium and lead. Mg had a significant (P < 0.01) correlation with K, Fe, Cu, and As. This study shows that honey bees (worker bees, drone bees, and bee broods) could be more useful as bioindicators of environmental element deposition (toxic and non-toxic) than multifloral honey. Also, this study shows that Serbian multifloral honey meets safety criteria concerning the concentrations of toxic elements.

Twenty-five-year study of Nosema spp. in honey bees (Apis mellifera) in Serbia.

A total of 7386 samples of adult honey bees from different areas of Serbia (fifteen regions and 79 municipalities) were selected for light microscopy analysis for Nosema species during 1992-2017. A selection of honey bee samples from colonies positive for microsporidian spores during 2009-2011, 2015 and 2017 were then subjected to molecular diagnosis by multiplex PCR using specific primers for a region of the 16S rRNA gene of Nosema species. The prevalence of microsporidian spore-positive bee colonies ranged between 14.4% in 2013 and 65.4% in 1992. PCR results show that Nosema ceranae is not the only Nosema species to infect honey bees in Serbia. Mixed N. apis/N. ceranae infections were detected in the two honey bee samples examined by mPCR during 2017. The beekeeping management of disease prevention, such as replacement of combs and queens and hygienic handling of colonies are useful in the prevention of Nosema infection.

Developing and Implementing Low-Cost Remote Laboratories for Undergraduate Biology and Neuroscience Courses.

The global pandemic caused by the novel coronavirus (SARS-COV-2) has forced many universities to abruptly change the delivery of courses from in-person to online. This change to remote learning requires creating new ways to deliver lectures, exams, and discussion groups through online meeting platforms. An often-overlooked challenge is performing lab courses that require access to specialized equipment and resources typically found in the undergraduate laboratory classrooms. Here we discuss some strategies for developing and implementing a full semester neuroscience laboratory course that allows students to fully participate in laboratory exercises at home or in their dorm rooms. Performing lab exercises remotely and independently was shown to significantly improve participant's self-efficacy and confidence that they can learn complex neuroscience material, when compared to participants who passively watch experiments online. We review best practices to ensure that lessons can be successfully demonstrated by the instructor and carried out by all students. Finally, we discuss the need to provide a level playing field such that all students may succeed, regardless of their current technology resources at home.

The Toxic Element Concentration in Fish Tissues from Sanicani Lake, an Urban Environment, in Bosnia and Herzegovina.

Concentrations of toxic elements (lead, cadmium, mercury, and arsenic) were measured in water, sediment, and four commercial fish species (Cyprinus carpio, Hypophthalmichthys molitrix, Silurus glanis, and Esox lucius) and their tissues (muscle, liver, and kidney) from Sanicani Lake, Bosnia and Herzegovina. The concentration of toxic elements was measured by atomic absorption spectrometry. Toxic element concentrations were significantly affected by the fish species and tissues sampled. The lead was the most heavily deposited element in lake sediments and waters. The highest concentrations of Pb and Cd were in the kidney in all four fish species. Concentrations of Hg and As were higher in the muscle than in the kidney and liver in all four fish species. The toxic element concentrations in all examined tissues were significantly higher in Silurus glanis and Esox lucius than in the other fish species examined. Concentrations of all tested elements were present in the fish muscle at different levels, but these levels were under the maximum levels allowed by the European Union and FAO/WHO. Accordingly, fish meat from this lake is acceptable for human consumption.

Novel application of the gray-level co-occurrence matrix analysis in the parvalbumin stained hippocampal gyrus dentatus in distinct rat models of Parkinson's disease.

To reveal the best choice of algorithm for parvalbumin-immunostained images of the hippocampal gyrus dentatus in two distinct rat models of Parkinson's disease (PD), particularly in terms of extracting the crucial information from the image, we tested whether the impact of experimentally induced dopaminergic (hemiparkinsonism) vs. cholinergic (PD cholinopathy) innervation impairment on the parvalbumin stained GABA interneurons could be detected using two separate algorithms, the fractal box-count and the gray-level co-occurrence matrix analysis (GLCM) algorithms. For the texture and fractal analysis of the hippocampal gyrus dentatus images, we used.tif images from three experimental groups of adult male Wistar rats: control rats, rats with Parkinson disease (PD) cholinergic neuropathology (with a PPT lesion), and hemiparkinsonian rats (with a SNpc lesion). For the suprapyramidal layer of the gyrus dentatus ASM and Entropy differentiated the images of the SNpc lesion versus the images of the control and the PPT lesion subjects, with significantly higher ASM and lower Entropy, indicating the homogenization of the images and their lower gray-level complexity. The infrapyramidal images of the SNpc group were differentiated versus the images from the control and PPT groups in terms of all the GLCM parameters: they showed lower mean Entropy and Contrast and higher ASM, Correlation and IDM. These results strongly suggest a rise in the uniformity, homogeneity and orderliness in the gray-levels of images from the SNpc group. Our results indicate that GLCM analysis is a more sensitive tool than fractal analysis for the detection of increased dendritic arborization in histological images.

Alterations of Sleep and Sleep Oscillations in the Hemiparkinsonian Rat.

Our previous studies in the rat model of Parkinson's disease (PD) cholinopathy demonstrated the sleep-related alterations in electroencephalographic (EEG) oscillations at the cortical and hippocampal levels, cortical drives, and sleep spindles (SSs) as the earliest functional biomarkers preceding hypokinesia. Our aim in this study was to follow the impact of a unilateral substantia nigra pars compacta (SNpc) lesion in rat on the cortical and hippocampal sleep architectures and their EEG microstructures, as well as the cortico-hippocampal synchronizations of EEG oscillations, and the SS and high voltage sleep spindle (HVS) dynamics during NREM and REM sleep. We performed unilateral SNpc lesions using two different concentrations/volumes of 6-hydroxydopamine (6-OHDA; 12 µg/1 µl or 12 µg/2 µl). Whereas the unilateral dopaminergic neuronal loss >50% throughout the overall SNpc rostro-caudal dimension prolonged the Wake state, with no change in the NREM or REM duration, there was a long-lasting theta amplitude augmentation across all sleep states in the motor cortex (MCx), but also in the CA1 hippocampus (Hipp) during both Wake and REM sleep. We demonstrate that SS are the hallmarks of NREM sleep, but that they also occur during REM sleep in the MCx and Hipp of the control rats. Whereas SS are always longer in REM vs. NREM sleep in both structures, they are consistently slower in the Hipp. The dopaminergic neuronal loss increased the density of SS in both structures and shortened them in the MCx during NREM sleep, without changing the intrinsic frequency. Conversely, HVS are the hallmarks of REM sleep in the control rats, slower in the Hipp vs. MCx, and the dopaminergic neuronal loss increased their density in the MCx, but shortened them more consistently in the Hipp during REM sleep. In addition, there was an altered synchronization of the EEG oscillations between the MCx and Hipp in different sleep states, particularly the theta and sigma coherences during REM sleep. We provide novel evidence for the importance of the SNpc dopaminergic innervation in sleep regulation, theta rhythm generation, and SS/HVS dynamics control. We suggest the importance of the underlying REM sleep regulatory substrate to HVS generation and duration and to the cortico-hippocampal synchronizations of EEG oscillations in hemiparkinsonian rats.

Sleep spindle dynamics during NREM and REM sleep following distinct general anaesthesia in control rats and in a rat model of Parkinson's disease cholinopathy.

On the basis of our previous studies and the important role of the thalamo-cortical network in states of unconsciousness, such as anaesthesia and sleep, and in sleep spindles generation, we investigated sleep spindles (SS) and high-voltage sleep spindle (HVS) dynamics during non-rapid eye movement (NREM) and rapid eye movement (REM) sleep following different types of general anaesthesia in both physiological controls and in a rat model of Parkinson's disease (PD) cholinopathy, to follow the impact of anaesthesia on post-anaesthesia sleep at the thalamo-cortical level through an altered sleep spindle dynamics. We recorded 6 hr of spontaneous sleep in all rats, both before and 48 hr after ketamine/diazepam or pentobarbital anaesthesia, and we used 1 hr of NREM or REM sleep from each to validate visually the automatically detected SS or HVS for their extraction and analysis. In the controls, SS occurred mainly during NREM, whereas HVS occurred only during REM sleep. Ketamine/diazepam anaesthesia promoted HVS, prolonged SS during NREM, induced HVS of increased frequency during REM, and increased SS/HVS densities during REM versus NREM sleep. Pentobarbital anaesthesia decreased the frequency of SS during NREM and the HVS density during REM sleep. Although the pedunculopontine tegmental nucleus lesion prolonged SS only during NREM sleep, in these rats, ketamine/diazepam anaesthesia suppressed HVS during both sleep states, whereas pentobarbital anaesthesia promoted HVS during REM sleep. The different impacts of two anaesthetic regimens on the thalamo-cortical regulatory network are expressed through their distinct sleep spindle generation and dynamics that are dependent on the NREM and REM state regulatory neuronal substrate.

Seasonal distributions of heavy metal concentrations in different snail (Helix pomatia) tissues from an urban environment in Serbia.

The aim of this study was to evaluate the differences between concentrations of heavy metals (cadmium, copper, zinc, iron, and manganese) in terrestrial gastropods, Helix pomatia, collected in four different environment locations in Pancevo city, Serbia. Metal concentrations in the foot and in the digestive gland were measured. Heavy metals cadmium, copper, zinc, iron, and manganese were measured in triplicate by atomic absorption spectrophotometry. Also, this study investigated correlations between the trace element content in the soft tissues and different seasons (summer and fall). The results showed that heavy metal concentrations of the measured elements in both snail tissues were significantly higher in polluted sites when compared to the background levels in the city. In digestive gland tissues, the concentration of heavy metals was higher compared with that in foot tissues. The bioaccumulation (heavy metal concentrations in soft tissues) of heavy metals (Cd, Cu, Zn, Fe, and Mn) in snails, Helix pomatia, has been assessed and related to seasons and sites. Heavy metal accumulation in the soft tissues of snails could provide a powerful monitoring tool for the assessment of environmental heavy metal pollution.

The effects of dietary Selenium-yeast level on glutathione peroxidase activity, tissue Selenium content, growth performance, and carcass and meat quality of broilers.

The present study was conducted to assess effects of selenium (Se)-yeast supplementation on glutathione peroxidase activity, Se levels in tissues, growth performance, carcass, and meat composition in broilers. A total of 275 one-d-old Cobb 500 broilers of both sexes were randomly allotted to 1 of 5 treatments during a 42-d period. The 5 treatments differed only in Se content: group 1 had no additional Se (background only); groups 2, 3, and 4 received 0.3 mg/kg of added Se from the beginning of the trial until d 21, whereas in the second half of the study (from d 22 to 42), these groups received 0.3, 0.6, and 0.9 mg/kg of added Se, respectively; and group 5 received 0.9 mg/kg of Se for the entire experimental period. At the end of the study, the control group showed significantly lower (P < 0.01) glutathione peroxidase activity in blood plasma compared to Se-supplemented groups. Regarding Se concentration in various tissues, the groups receiving Se yeast showed higher plasma, feces, and meat Se contents than the control group (P < 0.01). Supplementation of Se improved broilers' body weight, weight gain and feed conversion ratio (P < 0.01). Dressing percentage was lower in the control group and the group with 0.3 mg/kg of added Se compared to other experimental groups (0.6 and 0.9 mg/kg of dietary Se). The proportion of less valuable carcass parts (wings and legs) was higher (P < 0.01) in the group fed the basal diet compared to groups supplemented with 0.9 mg/kg of Se. Initial and ultimate pH values differed among experimental groups (P < 0.05). Supplementation of Se improved the broiler's antioxidative resistance, growth performance, carcass quality, and chemical composition of meat.

Physicochemical parameters and microbiological status of honey produced in an urban environment in Serbia.

Honey is a natural substance produced by honey bees (the genus Apis) enjoyed by people due to its unique nutritional and medicinal properties. The aim of this study was to determine the physicochemical parameters (moisture, ash, water-insoluble content, reducing sugars, sucrose, free acidity, diastase activity, hydroxymethylfurfural, and electrical conductivity) and microbiological status (total number of aerobic mesophilic bacteria, total number of sulfite-reducing clostridia, the presence of Salmonella spp., total numbers of fungi and yeasts and the presence of Clostridium botulinum) in honey (honeydew, blossom, sunflower, acacia, and linden) produced in an urban environment in Serbia. We analyzed 19 apiary samples of honey, collected during the 2011 harvesting season, by using recommendation methods. Physicochemical parameters of the examined honey produced in the urban environment indicated the honeys were of acceptable quality. Bacillus spp. were detected in four honeys, yeasts were detected in three honeys, and Clostridium botulinum type E was detected in one honey using PCR. The current study also showed the presence of diverse honey varieties in Serbia.

Effects of selenium source and level in diet on glutathione peroxidase activity, tissue selenium distribution, and growth performance in poultry.

Today, a few differing sources of selenium (Se), i.e. inorganic, organic, and nano forms of Se, are used as feed supplements for poultry. Published research indicates that nano-Se and organic Se possess comparable efficiency to inorganic Se in increasing GSH-Px activity of plasma and various tissues, but they deposit at higher rates in various tissues. However, there are principal differences in absorption mechanisms, metabolism, and efficiency of these three forms of Se. The aim of this review was to analyze the available literature on the effects of different Se sources and levels in the diet on glutathione peroxidase (GSH-Px) activity, tissue Se distribution and growth performance in poultry. Higher levels of Se increase GSH-Px activity in the body, but this reaches a plateau even if Se concentrations in diet increase further, while the deposition of Se in tissues increases as Se content in diet increases. In addition, many studies have shown the positive effects of adding Se to diet on growth performance in poultry. Optimal Se supplementation is necessary not only for good poultry health but also to ensure and preserve meat quality during storage and to provide human beings with this microelement.

Sleep disorder and altered locomotor activity as biomarkers of the Parkinson's disease cholinopathy in rat.

In order to find out the possible earliest biomarkers of Parkinson's disease (PD) cholinopathy, we followed the impact of bilateral pedunculopontine tegmental nucleus (PPT) lesion in rat on: the cortical and hippocampal sleep/wake states architectures, all sleep states related EEG microstructures, sleep spindles, the basal and stimulated locomotor activity. Sleep and basal locomotor activity in adult Wistar rats were followed during their inactive circadian phase, and throughout the same aging period. The bilateral PPT lesions were done by 0.1M ibotenic acid (IBO) during the surgical procedure for implantation of the electroencephalographic (EEG) and electromyographic (EMG) electrodes for chronic sleep recording. The cholinergic neuronal loss was identified by NADPH - diaphorase histochemistry. After all sleep and behavioral recording sessions, the locomotor activity was stimulated by d-amphetamine (d-AMPH) and the neuronal activity of striatum was followed by c-Fos immunolabeling. Impaired cholinergic innervation from the PPT was expressed earlier as sleep disorder then as movement disorder, and it was the earliest and long-lasting at hippocampal and thalamo-cortical level, and followed by a delayed "hypokinesia". This severe impact of a tonically impaired PPT cholinergic innervation was evidenced as the cholinergic interneuronal loss of the caudate putamen and as a suppressed c-Fos expression after stimulation by d-AMPH. In order how they occurred, the hippocampal non rapid eye movement (NREM) sleep disorder, altered high voltage sleep spindle (HVS) dynamics during rapid eye movement (REM) sleep in the hippocampus and motor cortex, and "hypokinesia" may serve as the biomarkers of PD cholinopathy onset and progression.

Determination of heavy metals in muscle tissue of six fish species with different feeding habits from the Danube River, Belgrade-public health and environmental risk assessment.

The aim of this study was to investigate the presence of heavy metals/metalloids (Pb, Cd, Hg, Cu, Fe, Zn, As) in the muscle tissue of fish from the Danube River (two locations: Zemun and Grocka). For the purpose of heavy metal determination in fish muscle, 120 samples of six different fish species, Prussian carp, barbel, bream, carp, pike perch, and catfish were collected. For determining heavy metals, we used microwave oven digestion and atomic absorption spectrometer methods. The highest average content of Pb (0.084 ± 0.004 mg kg-1), Cd (0.082 ± 0.003 mg kg-1), Hg (0.466 ± 0.006 mg kg-1), and As (0.333 ± 0.007 mg kg-1) was found in the muscle of carp (an omnivorous fish) from Grocka, while the highest average level of Fe (13.60 ± 0.03 mg kg-1) was deposited in bream (also omnivorous) from Zemun. Also, the average Cu level (1.62 ± 0.13 mg kg-1) was the highest in catfish muscle (a carnivorous fish) from Grocka, while the highest Zn content (11.16 ± 0.17 mg kg-1) was determined in muscle of Prussian carp (an omnivorous fish) from Zemun. The highest content of heavy metals (Cu, Fe, and Zn, respectively) in muscle of the six different types of fish from both locations was symmetrically arranged by species (catfish, barbel, and Prussian carp, respectively). Concentrations of Pb, Hg, and As in the Danube River fish muscle were under the maximum residual levels prescribed by the European Union (EU) and the maximum allowed concentrations (MAC) for Serbia. On the other hand, in all fish muscle from both locations (Zemun and Grocka), higher concentrations of Cd than prescribed (MAC) were found, with the exception of bream and pike perch.