Humic Substances as a Versatile Intermediary.

Humic substances are organic ubiquitous components arising in the process of chemical and microbiological oxidation, generally called humification, the second largest process of the carbon cycle. The beneficial properties of these various substances can be observed in many fields of life and health, whether it is the impact on the human organism, as prophylactic as well as the therapeutic effects; animal physiology and welfare, which is widely used in livestock farming; or the impact of humic substances on the environment and ecosystem in the context of renewal, fertilization and detoxification. Since animal health, human health and environmental health are interconnected and mutually influencing, this work brings insight into the excellence of the use of humic substances as a versatile mediator contributing to the promotion of One Health.

Humic Substances as a Feed Supplement and the Benefits of Produced Chicken Meat.

Humic substances with a high proportion of humic acids (more than 40%) have been classified by the European Commission as feed materials that can be used in animal nutrition since 2013. A protective effect on the intestinal mucosa, as well as anti-inflammatory, adsorptive and antimicrobial properties, were recorded. Nutrient absorption, nutritional status and the immune response in chickens supplemented with HSs were significantly improved. HSs have the ability to enhance protein digestion as well as the utilization of calcium and trace elements. They are known to improve feed digestibility as a result of maintaining an optimal pH in the gut, leading to lower levels of nitrogen excretion and less odor in the husbandry environment. HSs not only increase digestibility and result in greater utilization of the feed ration but also improve the overall quality of the meat produced. They increase the protein content and reduce the fat content in breast muscles. They also contribute to improving the sensory characteristics of the meat produced. Their antioxidant properties improve the oxidative stability of meat during storage. The influence of HSs on fatty acid composition may be one of the reasons that meat has a more beneficial effect on the health of consumers.

The Effect of the Supplementation of Humic Substances and Fermented Products in the Feed on the Content of Salinomycin Residues in Poultry Tissues.

The presence of antimicrobial residues in products of animal origin is a constant problem for consumer health. The aim of this study was to observe the effect of the addition of humic substances (H), fermented products (F) and a mixture of both (FH) to feed supplemented with the coccidiostat salinomycin, compared with a control group (C), on the content of salinomycin residues in the edible tissues of broiler chickens using two microbial inhibition screening methods, Explorer 2.0 test and the Screening Test for Antibiotic Residues (STAR), and a confirmatory competitive enzyme immunoassay analysis (Salinomycin ELISA Kit). The results of the microbial inhibition tests showed a gradual decline in the positive results in the tissue samples from the last day of salinomycin administration (30th day) tothe last day of fattening (37th day, day of slaughter) in group C and no positive results in the tissue samples from experimental groups H, F and FH slaughtered on the last day of fattening. Using the Salinomycin ELISA Kit, salinomycin was detected in the chicken muscle tissues of all the control and experimental groups. However, no sample from any group contained salinomycin at a concentration exceeding the maximum residue limits set by European law. The high level of significance (p < 0.001) confirmed the positive influence of the administration of humic substances and fermented products on the content of salinomycin residues in chicken tissues.

Detection of Microbiota during the Fermentation Process of Wine in Relation to the Biogenic Amine Content.

Viticulture is one of the traditional industries in Slovakia, where there are six wine-growing regions: Malokarpatska, Southern Slovakia, Central Slovakia, Nitra, Eastern Slovakia, and Tokaj. This study focuses on the detection of microbiota in soil samples, grape leaves and berries, and samples taken from fermenting must and young wine (the variety Tramín cervený) in relation to the detected concentrations of biogenic amines during the fermentation process. In the examined samples, the number of yeasts and molds (from 3.8 to 6.8 log cfu/g or mL) and TVC (from 3.7 to 6.5 log cfu/g or mL) were determined via culture examination. At the same time, the number of LAB (from ˂3.0 to 4.4 log cfu/g or mL) was determined, which was the highest on day 4 of the must fermentation process and was related to the detected of the highest concentration of biogenic amines (histamine and tyramine) on day 6 in the investigated must samples using the UHPLC system. Mycobiota species were identified by MALDI-TOF MS, PCR, ITS-PCR-RFLP, and PCR sequencing of the amplified products. The study confirmed the presence of the yeasts Saccharomyces cerevisiae, Metschnikowia pulcherrima, Hanseniospora uvarum, Pichia kudriavzevii, Pichia kluyveri, Pichia fermentas, Torulaspora delbrueckii, and Candida tenuis. At the same time, the presence of molds (Cladosporium herbarum, Cladosporium cladosporioides, Penicillium granulatum, Penicillium mononematosum, Botritis cinerea, and Penicillium glabrum) was also confirmed in soil samples, leaves, grape berries, and fresh grape must. The study confirmed the reduction in the species diversity of the microbiota during the must fermentation process, which resulted in decreases in the concentrations of the monitored biogenic amines in the early stages of the must fermentation process and young wine of the variety Tramín cervený.

Detection of Gluten in Gluten-Free Foods of Plant Origin.

The work deals with the issue of standardization and more accurate methodology for the isolation of gluten DNA in gluten-free products of plant origin, which is more demanding due to the more complex structure of plant cells. Three isolation methods were compared, of which the combination of glass and zirconium beads, Proteinase K and a commercially produced isolation kit was confirmed to be the most effective procedure. The given isolation procedure was more effective in one-component gluten-free foods, where the concentration of the obtained DNA ranged from 80.4 ± 0.7 to 99.0 ± 0.0 ng/µL. The subsequent PCR reaction revealed the presence of gluten not only in guaranteed gluten-free products (40%), but also in naturally gluten-free foods (50%). These were mainly gluten-free sponge cakes, gluten-free biscuits "Cranberries", cocoa powder, coffee "3in1", and instant coffee.

Effect of Concentration of Flaxseed (Linum usitatissimum) and Duration of Administration on Fatty Acid Profile, and Oxidative Stability of Pork Meat.

Flaxseed is a common ingredient used for livestock feed. The aim of this work was to study the effect of a diet supplemented with flaxseed at 5% and 10% concentrations in the intervals of 3 and 6 weeks prior slaughter on fatty acid profile and oxidative stability of pork meat. Meat samples were collected after slaughter from each animal (five groups, n = 6). Samples of the musculus longissimus dorsi (MLD) and the musculus gluteobiceps (MGB) were selected. Chemical composition, fatty acid profile and oxidative stability during the storage of meat under chilling conditions (4 °C, 7 days) was analyzed. The addition of flaxseed significantly affected the composition of fatty acid profile and the shelf life of the produced meat. The fat content was changed in the experimental groups with 10% flaxseed supplementation (10.84% in MGB and 9.56% MLD) versus the control group. Despite the different concentrations of flaxseed, the best EPA/AA ratio was observed in the experimental groups fed with flaxseed supplementation for 3 weeks. The worst oxidative stability of meat samples (p < 0.05) was recorded in the experimental groups with the addition of flaxseed for 6 weeks, which was related to higher PUFA content in samples of the experimental groups and higher susceptibility of PUFAs to lipid oxidation. The oxidative stability of meat in the experimental group fed 5% flaxseed supplementation for 3 weeks was not affected.

Antimicrobial Resistance of Enterococcus sp. Isolated from Sheep and Goat Cheeses.

This study aimed to calculate the proportion of antibiotic resistance profiles of Enterococcus faecium, E. faecalis, and E. durans isolated from traditional sheep and goat cheeses obtained from a selected border area of Slovakia with Hungary (region Slanské vrchy). A total of 110 Enterococcus sp. were isolated from cheese samples, of which 52 strains (E. faecium (12), E. faecalis (28), E. durans (12)) were represented. After isolation and identification by polymerase chain reaction and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry, the enterococci (E. faecium, E. faecalis, and E. durans) were submitted to susceptibility tests against nine antimicrobial agents. In general, strains of E. faecalis were more resistant than E. durans and E. faecium. A high percentage of resistance was noted in E. faecalis to rifampicin (100%), vancomycin (85.7%), teicoplanin (71.4%), erythromycin (71.4%), minocycline (57.1%), nitrofurantoin (57.1%), ciprofloxacin (14.3%), and levofloxacin (14.3%). E. durans showed resistance to rifampicin (100%), teicoplanin (100%), vancomycin (66.7%), erythromycin (66.7%), nitrofurantoin (66.7%), and minocycline (33.3%), and E. faecium showed resistance to vancomycin, teicoplanin, and erythromycin (100%). Multidrug-resistant strains were confirmed in 80% of the 52 strains in this study. Continuous identification of Enterococcus sp. and monitoring of their incidence and emerging antibiotic resistance is important in order to prevent a potential risk to public health caused by the contamination of milk and other dairy products, such as cheeses, made on farm level.

Heavy-Metal Contents and the Impact of Roasting on Polyphenols, Caffeine, and Acrylamide in Specialty Coffee Beans.

The aim of this study was to determine the effect of roasting on the contents of polyphenols (PPH), acrylamide (AA), and caffeine (CAF) and to analyze heavy metals in specialty coffee beans from Colombia (COL) and Nicaragua (NIC). Samples of NIC were naturally processed and COL was fermented anaerobically. Green beans from COL (COL-GR) and NIC (NIC-GR) were roasted at two levels, light roasting (COL-LIGHT and NIC-LIGHT) and darker roasting (COL-DARK and NIC-DARK), at final temperatures of 210 °C (10 min) and 215 °C (12 min), respectively. Quantitative analyses of PPH identified caffeoylquinic acids (CQA), feruloylquinic acids, and dicaffeoylquinic acids. Isomer 5-CQA was present at the highest levels and reached 60.8 and 57.7% in COL-GR and NIC-GR, 23.4 and 29.3% in COL-LIGHT and NIC-LIGHT, and 18 and 24.2% in COL-DARK and NIC-DARK, respectively, of the total PPH. The total PPH contents were highest in COL-GR (59.76 mg/g dry matter, DM). Roasting affected the contents of PPH, CAF, and AA (p < 0.001, p < 0.011 and p < 0.001, respectively). Nickel and cadmium contents were significantly higher in the COL-GR than in the NIC-GR beans. Darker roasting decreased AA content, but light roasting maintained similar amounts of CAF and total PPH.

Effect of Broilers Chicken Diet Supplementation with Natural and Acidified Humic Substances on Quality of Produced Breast Meat.

This study was conducted to examine the effect of two humic substances (HS) supplemented in broilers' diet on the breast meat quality of broiler chickens. In this experiment, 120 pieces of one-day-old COBB500 broiler chickens were used. Broilers were divided into three groups, each containing 40 birds with three replications (13, 13, and 14 per one pen). Fattening lasted 38 days. The first experimental diet was supplemented with 0.7% of HS (HS0.7) and the second was enriched with 0.7% of acidified HS (HSA0.7). The control group of broilers (C) was fed a basal diet without the addition of any supplements. HS0.7 samples had the highest total protein content and the lowest content of fat (p < 0.01). The effects of broiler diet and storage had a significant impact on the pH of breast samples, p < 0.001 and p < 0.05, respectively. Supplementation of HS in broiler diet positively affected the lipid oxidation of breast meat samples, as indicated by the lower malondialdehyde content (p < 0.05). HSA0.7 samples had significantly better juiciness after the storage period (p < 0.001). The quality of meat produced with supplementation of HS in broilers' feed can be considered as valuable in human nutrition due to improved protein and fat content with decreased lipid oxidation.

Detection of Carrageenan in Meat Products Using Lectin Histochemistry.

Carrageenan is a polysaccharide that is widely used in the food industry. Due to its water holding capacity, there is a higher risk of adulteration for economic reasons related to it. A verifiable method for detecting carrageenan is still missing in the food inspection sector. The detection of carrageenan in meat products is not well described. Our study describes lectin histochemistry as a novel approach for carrageenan detection. Within this study, the detection of carrageenan in meat products by lectin histochemistry is validated. Lectins of Arachis hypogaea (PNA) and Bandeiraea simlicifolia (BSA), specific for galactose units of carrageenan, were used. The samples included model meat products (ground chicken-meat products) and meat products from retail markets (chicken and pork hams, sausages, salami, and dried sausages). The limit of determination (LoD) of this method was set at 0.01 g kg-1. The method sensitivity for lectin PNA reached 1, and, for lectin BSA, it reached 0.96. Method specificity for lectin PNA was 1, and, for lectin BSA, it was 1.33. Cross-reactivity with other hydrocolloids tested was not confirmed. The results confirm that lectin histochemistry is suitable for detecting carrageenan in meat products.

Effect of Solid-State Fermented Wheat Bran Supplemented with Agrimony Extract on Growth Performance, Fatty Acid Profile, and Meat Quality of Broiler Chickens.

The impact of the broiler diet modification on the following parameters was evaluated: meat quality, carcass traits, and bone and blood parameters. One hundred twenty one-day-old COBB 500 broiler chickens were assigned to three experimental groups (40 birds per group) with four replications (10 per pen) for 35 days of fattening. The control (C) was fed a basic feed mixture. The diet supplemented with 10% of fermented feed (FF10) and additionally supported by 0.2% of agrimony extract (FF10 + AE) was applied to the second and third groups, respectively. FF10 showed both a lower average daily feed intake and total feed consumption when compared to that of C (p < 0.05). Lower concentration of alkaline-phosphatase and calcium and higher total lipids and triglycerides in blood were observed in FF10 + AE. Breast and thigh meat showed a lower content of polyunsaturated fatty acid n-3 and n-6 in the FF10 + AE group (p < 0.01). The increase of gamma-linolenic acid content in breast and thigh meat samples obtained from the experimental groups was significant (p < 0.001 and p < 0.05; respectively). Lower lipid oxidation was observed in the thigh muscle of the FF10 + AE group on the first day of storage (p < 0.01). The current study indicates that FF10 + AE supplementation can be successfully applied to enhance broiler performance and meat quality.

Multiple factorial analysis of physicochemical and organoleptic properties of breast and thigh meat of broilers fed a diet supplemented with humic substances.

Multiple factor analysis was used for the examination of meat quality of broilers, the diet of which was supplemented with 0.8 and 1.0% addition of humic substances (HS). One hundred fifty COBB 500 one-day-old male broilers chicks were randomly divided into 3 different groups: one control and 2 experimental (n = 50), and they were fattened for 35 D. Subsequently, the meat quality was analyzed and defined by physicochemical and sensory quality, supplemented with analysis by instrumental methods. We observed changes in dry matter, fat, water, and protein content in experimental samples of breast and thigh meat (P < 0.001). In both experimental groups, the concentration of phosphates and pH decreased in breast meat (P < 0.001) and in thigh meat (P < 0.05). The smell of experimental chicken breast meat samples after cooking was evaluated by a sensory panel, which scored a higher point score than that of the control group (P < 0.05). Sensory evaluation of taste indicated a positive response with respect to the perception of meat quality in relation to a greater supplementation of HS in the diet. Thigh meat samples showed a variable extent of water loss after cooking, but lower values of water loss were generally obtained from thigh meat samples of poultry fed with higher HS supplementation, than in chicken breast meat samples. Significant differences in evaluated variables between both experimental groups were not observed. The color of breast meat samples changed, when considering the variables of lightness and redness, with the addition of 1% HS (P < 0.05). The main advantage of the breast meat of broilers fed a diet supplemented with HS was observed in the final meat quality, which was positively affected by increased protein and decreased fat content. Because of its nutritional composition, it can be considered to be rather a valuable type of meat in human nutrition than ordinarily.

Dual production of polyunsaturated fatty acids and beta-carotene with Mucor wosnessenskii by the process of solid-state fermentation using agro-industrial waste.

Solid-state fermentation is a technique employing microorganisms grown on a solid substrate in the absence of free water. The substrates used in this process are mostly waste from the agro-industry (brans, spent malt grains, distiller grains, etc.) that improves not only the economy of the process but also has positive effect on waste management problems. Zygomycetous fungi are not only able to grow in such conditions but also enrich fermented materials with various types of bioactive compounds. Mucor sp. strains have been identified as producers of gamma-linolenic acid and beta-carotene in submerged fermentation. The aim of the present study was to identify the best microbial producer of gamma-linolenic acid and beta-carotene among four different Mucor strains and to study the requirements for the dual production of these metabolites. Mucor wosnessenskii was identified as the most suitable producer of both metabolites. After optimization of the fermentation conditions, the highest yields obtained were 10.7 g of gamma-linolenic acid/kg of fermented product and 261.5 mg of beta-carotene/kg of fermented product. This yield of beta-carotene is the highest among the results published so far.

Effect of Fungal Solid-State Fermented Product in Broiler Chicken Nutrition on Quality and Safety of Produced Breast Meat.

The aim of this work was to analyse the effect of addition of 10% (w/w) fermented bioproduct into commercial broiler feed on fatty acid profile, lipid oxidative stability, and sensory properties of chicken breast meat. The fermented bioproduct was prepared by fermentation of cornmeal by filamentous fungi Umbelopsis isabellina CCF 2412 in solid-state fermentation (SSF) process and the final bioproduct was enriched with gamma-linolenic acid and beta-carotene. In the experiment, 80 pieces of 1-day-old chickens COBB 500 were used. Half of them (control group) were fed only with commercial feed. Chickens of the experimental group were fed with commercial feed, and, from the 11th day of age until the time of slaughter (39th day), 10% of commercial feed was replaced with fermented bioproduct. Application of fermented bioproduct into commercial feed mixture positively influenced profile of fatty acids in breast meat. The amount of gamma-linolenic, alpha-linolenic, and oleic acids in fat of breast muscles was increased and n-6/n-3 ratio was significantly decreased. Profile and content of PUFAs did not change after thermal treatment of meat. Oxidative stability of fat and sensory properties of the meat during the storage (4°C, 7 days) of meat were not affected by fermented bioproduct.

Chlamydiosis in farmed chickens in Slovakia and zoonotic risk for humans.

INTRODUCTION: Chlamydia psittaci is an obligate intracellular Gram-negative bacterium causing respiratory disease (chlamydiosis) or asymptomatic carriage in poultry. In humans, it is a zoonotic agent of ornithosis/psittacosis. Due to low awareness of the disease and variable clinical presentation, psittacosis is often remains unrecognised as such by general practitioners. Zoonotic transfer occurs through inhalation of contaminated aerosols, and originates from feathers, faecal material and respiratory tract exudates. OBJECTIVE: The aim of this study was to investigate chickens for the presence of Chlamydia sp. from pharyngeal and cloacal swabs and review the zoonotic risk for humans. MATERIAL AND METHODS: 138 clinically healthy chickens from farms in Slovakia were examined for the presence of Chlamydia sp. The age of the chickens was 6 months. Two different samples were used - pharyngeal swabs and cloacal swabs. Each sample was examined by the molecular PCR method, and in the case of a positive result the identity of the obtained sequences was examined by a BLAST search. RESULTS: Of the total number of 276 examined samples from 138 chickens, 19 (6.9%) showed positivity for C. psittaci infection, 12 (8.7%) which were positive from pharyngeal swabs and 7 (5.1%) from cloacal swabs. None of the chickens were positive in both samples. Phylogenetic examination of the 19 isolates identified in the study, based on the 23S rRNA gene sequence, revealed that the isolates obtained were identical with C. psittaci, and genetically very close to genotypes B and genotype E. CONCLUSIONS: C. psittaci infections are apparently emerging in chickens. Chicken-processing plant employees should be considered a risk group for human psittacosis. There is a need for higher awareness and for efficient risk assessment and management.

Lipid Peroxidation Process in Meat and Meat Products: A Comparison Study of Malondialdehyde Determination between Modified 2-Thiobarbituric Acid Spectrophotometric Method and Reverse-Phase High-Performance Liquid Chromatography.

The aim of this work was to compare the methods of malondialdehyde detection, as the main secondary product of the lipid peroxidation process, in meat and meat products. Malondialdehyde measurements were performed by two modified methods, the 2-thiobarbituric acid spectrophotometric method and the reverse-phase high-performance liquid chromatography in raw, mechanically-deboned chicken meat and in manufactured frankfurters. The malondialdehyde concentrations measured by the 2-thiobarbituric acid spectrophotometric method were found to be overestimated by more than 25% in raw meat and more than 27% in frankfurters in comparison to the results of reverse-phase high-performance liquid chromatography (p < 0.05). The achieved results showed that the presented modified reverse-phase high-performance liquid chromatography method was more applicable and more accurate for the quantification of malondialdehyde in samples of meat and meat products.

Effects of vacuum packaging conditions on the quality, biochemical changes and the durability of ostrich meat.

In this study, the effects of vacuum packaging conditions on the concentrations of lactic acid, malondialdehyde, pH, microbial and sensory analysis were determined during chilled storage of ostrich meat. Meat was packed as follows: vacuum packed from 1st day (VP-1), vacuum packed from 3rd day (VP-3) and non-vacuum packed (NVP). Analysis were performed at 1st, 7th, 14th, 21st day after slaughter. Meat consisted of 74.69% water, 2.29% fats, 20.95% proteins. Package conditions had significant effect on the pH (NVP: 6.54 on the 14th day, VP-1: 6.05 and VP-3: 6.07 on the 21th day p<0.001), amount of malondialdehyde (NVP: 8.62mg/kg on the 14th day; VP-1: 1.95 and VP-3: 2.55 on the 21th day; p<0.001) and total microbial count (NVP: 7.4 log CFU/g on the 14th day; VP-1: 6.7 and VP-3: 6.8 on the 21th day p<0.01). Based on these results we can assess that vacuum packed from 1st day is necessary for the 21 days storage of ostrich meat.

Post-mortem changes in the concentration of lactic acid, phosphates and pH in the muscles of wild rabbits (Oryctolagus cuniculus) according to the perimortal situation.

In this study changes in the concentrations of lactate, phosphates, and pH values of water extracts of muscles of transported and hunted rabbits during ripening were determined. Concentrations of lactate were higher in the muscles of hunted rabbits. The highest differences were obtained 24h after kill/hunt. Concentrations of lactate in the muscles of hunted rabbits were decreasing, while in the muscles of transported rabbits we observed it to increase in the 7th day and then decrease in the 14th day. Higher concentrations of phosphates were found in the muscles of transported wild rabbits. During the ripening process concentrations of phosphates were decreasing in muscles of both groups. Muscles of hunted rabbits had lower pH values during the whole ripening process. Our research showed that concentrations of lactate, phosphates and pH value post-mortem depended on the perimortal situations.

Determination of lipid oxidation level in broiler meat by liquid chromatography.

An assay was conducted for the determination of malondialdehyde (MDA) levels in broiler meat. The method involves extraction of tissues with trichloroacetic acid (TCA) and reaction of the TCA extract with 2,4-dinitrophenylhydrazine (DNPH). After separation of the MDA-DNPH complex using a solid-phase extraction C18 column, samples were eluted with 1 mL acetonitrile. Aliquots of 20 microL acetonitrile were analyzed by liquid chromatography on reversed-phase C18 column (3 microm) with UV detection. The products were eluted isocratically with the mobile phase containing acetonitrile-water-acetic acid (39 + 61 + 0.2, v/v/v). The retention time for MDA-DNPH was 6.5 min, and the detection limit was 3.5 microg/kg. Two extraction methods (cold and hot) were also used in the study. The results showed that hot extraction increased results about 55.8% and recovery from samples spiked with 116.6 microg/kg was lower (74.6%) in comparison with cold extraction (94.7%).