Iveta Plachá

Pathogens and antibiotic residues in animal manures and hygienic and ecological risks related to subsequent land application

The practice of spreading of livestock wastes onto land used for the production of food or animal feeds is widely regarded as the least environmentally damaging disposal method, however, the practice is still fraught with pitfalls such as N pollution of air and water and significant microbiological risks. Therefore this paper focuses on some of the latest developments that provide new insights into the microbiological safety of animal manures, the related treatment options and the spreading the products onto land. In conclusion the paper stresses the need to fully address issues concerning environmental contamination and transmission of antimicrobial-resistant bacteria through livestock manure, improve current environmental regulations regarding manure management practice and coordination of research activities and dissemination of technical information.

Hygienic and ecological risks connected with utilization of animal manures and biosolids in agriculture

Abstract In recent years the fate of human and animal pathogen microorganisms as a potential pollutant of the environment has been paid increased attention. Substantial quantities of these compounds and their metabolites are excreted, flushed down the drain, discarded as waste, or left over in animal feedlots. After passing to the sewer, several of these compounds are not adequately eliminated by the methods that are currently used in sewage treatment. Substantial quantities of biosolids and livestock manure end up on agricultural land. Effective sanitation of the environment, particularly of some of its special parts, which can be a source of spreading of diseases, plays an important role in prevention of infectious diseases. In this respect special attention should be paid to the disinfection of infected farm animal excrements. Sanitation of excrements should, on the one hand, ensure effective inhibition of infectious agents and, on the other hand, comply with the requirement of preserving the composition of the manure so it can be used in agricultural production.

Effects of deoxynivalenol and zearalenone on oxidative stress and blood phagocytic activity in broilers

Effects of dietary contamination with various levels of deoxynivalenol (DON) and zearalenone (ZEA) were investigated on Ross 308 hybrid broilers of both sexes. After hatching, all chickens were fed an identical control diet for two weeks. Then chickens of Group 1 received a diet contaminated with DON and ZEA, both being 3.4 mg · kg−1, while Group 2 received DON and ZEA at 8.2 and 8.3 mg · kg−1, respectively. The diet of the control group contained background levels of mycotoxins. Samples of blood and tissues were collected after two weeks. Intake of both contaminated diets resulted in a significantly decreased activity of glutathione peroxidase (GPx) and increased level of malondialdehyde (MDA) in liver tissue, while in kidneys the concentration of MDA was significantly increased only in Group 1. On the other hand, activities of blood GPx and plasma γ-glutamyltransferase (GGT) were elevated in Group 2 only. Activities of thioredoxin reductase in liver and GPx in duodenal mucosa tissues, superoxide dismutase (SOD) in erythrocytes as well as levels of MDA in duodenal mucosa and α-tocopherol in plasma were not affected by dietary mycotoxins. Blood phagocytic activity was significantly depressed in Group 1 and 2. These results demonstrate that diets contaminated with DON and ZEA at medium levels are already able to induce oxidative stress and compromise the blood phagocytic activity in fattening chickens.

Effects of excessive selenium supplementation to diet contaminated with deoxynivalenol on blood phagocytic activity and antioxidative status of broilers.

This study examined the effects of excessive dietary supplementation with organic selenium on phagocytic activity and antioxidative status of chickens for fattening fed diet contaminated with deoxynivalenol (DON). Sixty chickens of Ross 308 hybrids were at day of hatching divided into four groups with 15 birds in each. The background DON dietary levels in both negative and positive control groups were 0.2 mg/kg. The complete feed for positive control group was supplemented with Se dose 1 mg/kg in the form of Se-yeast. Group 3 was fed diet with DON level 3 mg/kg while diet for group 4 combined DON level 3 mg/kg with a excessive supplement of Se-yeast (Se dose 1 mg/kg). After 6 weeks of dietary intake, six randomly-chosen chickens from each group were sampled. Feeding of contaminated diet resulted in significantly reduced blood phagocytic activity (19.5 ± 1.1% in the negative control vs. 12.8 ± 0.8% in the DON-treated group, p < 0.05). Se-yeast supplemented to the DON contaminated diet prevented suppression of phagocytic activity. Dietary intake of DON at levels 3 mg/kg did not influence the plasma α-tocopherol level while excessive dietary Se dose reduced it in both Se supplemented groups. Neither the birds of DON-treated group nor the birds from group 4 with DON and Se-yeast showed any response in plasma γ-glutamyltransferase (GGT) activity. Subtoxic dietary level of DON significantly increased the activity of glutathione peroxidase (GPx) in the duodenal mucosa, while additional Se supplementation prevented such a response to mycotoxin. On the other hand, both Se supplemented groups showed significantly elevated GPx activities in blood, liver and kidney, (p < 0.05). The results suggest a potential ability of excessive supplementation of organic selenium to prevent the blood phagocytic activity suppression and changes in GPx activity in duodenal tissue induced in broilers by subtoxic dietary levels of DON.

Effect of Enterococcus faecium AL41 and Thymus vulgaris essential oil on small intestine integrity and antioxidative status of laying hens.

We investigated the effect of Enterococcus faecium on phagocytic activity, antioxidative status in vivo and the effect of E. faecium and 0.4% concentration of Thymus vulgaris essential oil (EO) on the duodenal tissue integrity in vitro in laying hens. The birds were fed the same standard diets and were divided into four groups. E. faecium was added to the drinking water for the second and fourth groups. EO was added to special chambers for measuring trans-epithelial electrical resistance (TEER) for the third and fourth groups only. TEER was lower in groups where EO was added, but in the group with E. faecium TEER was not changed significantly. Our results show that EO at 0.4% concentration may negatively affect intestine integrity, and the probiotic strain E. faecium AL41 is able to eliminate this effect and can strengthen non-specific immunity. To confirm our findings further histopathological investigations of intestinal tissue are needed.

Effect of thyme oil on small intestine integrity and antioxidant status, phagocytic activity and gastrointestinal microbiota in rabbits.

The effects of 0.5 g thyme oil per kg dry matter (DM) of diet on duodenal tissue integrity, antioxidant status, phagocytic activity and selected microbiota in the caecum and faeces of rabbits were studied. Twenty-four rabbits were divided into two groups and were fed a commercial granulated diet for growing rabbits (CD) with access to water ad libitum. The first group was fed the CD, while to the CD of the second group thyme oil was added. Intestinal integrity was tested by measuring the transepithelial electrical resistance (TEER). Thyme oil significantly increased the value of total antioxidant status (TAS) in the blood plasma and glutathione peroxidase (GPx) activity in the liver, and it decreased malondialdehyde (MDA) concentration in the duodenal tissue. Thyme oil resulted in strengthened intestinal integrity, as the essential oil supplementation significantly increased TEER values in the experiment. The faecal microbiota of rabbits was almost completely balanced in both groups, and only a slight decrease was found in the microbial population at day 42 of the trial. In both groups, the bacterial counts were generally lower in the caecum than in the faecal samples. In conclusion, dietary supplementation with 0.5 g/kg DM thyme oil may improve intestinal integrity, and it may have an antioxidant effect. A tendency was also found for thyme oil to stimulate the abundance of some microbes beneficial in the rabbit gut.

Can enterocins affect phagocytosis and glutathione-peroxidase in rabbits?

The gastrointestinal microbiota in rabbits play an important role in protection against potential pathogens via the development of the mucosal immune system. The gut health, including the microbial and immunological stability, is often influenced by exogenous factors, mainly around the weaning period. Therefore, alternative strategies are required to improve the animal’s health. In this study, the diet of rabbits was supplemented with the semi-purified enterocins Ent 2019, Ent M and Ent 4231, produced by bacteriocinogenic strains with probiotic properties, rabbit-derived Enterococcus faecium CCM7420 and non rabbit-derived E. faecium strains AL41 and CCM4231. The phagocytic activity, index of phagocytic activity and the gluthatione-peroxidase enzyme activity in blood were determined during the Ents consumption and also 3 weeks after their cessation. At 21 days into the experiment the highest phagocytic activity was observed in rabbits receiving Ent M. A significant increase in phagocytosis was noted in rabbits with Ent 2019 over the entire experiment. Moreover, a lower gluthatione-peroxidase activity was measured in rabbits receiving Ent M and Ent 2019. No effect of Ent 4231 application on the tested parameters was recorded. The Ent M and Ent 2019 improved the digestive immunity and the host’s defense capacities by stimulating leucocyte phagocytosis, without oxidative stress induction in rabbits.

Effect of lignin on oxidative stress in chickens fed a diet contaminated with zearalenone

The effect of lignin supplementation to a diet contaminated with zearalenone (ZEA) on antioxidant status was studied in female chickens of the ISA BROWN laying strain. From the day of hatching to 2 weeks of age, four groups of chickens were fed the same uncontaminated control diet. After 14 days, Group 1 (control) continued to receive the uncontaminated diet, while Group 2 was fed an identical diet enriched with 0.5% chemically modified lignin. Simultaneously, chickens of Group 3 were switched to a diet contaminated with 7.9 mg/kg ZEA and those of Group 4 to an identical contaminated diet supplemented with 0.5% lignin. At 6 weeks of age blood and tissue samples were collected. Feeding of a diet contaminated with a high level of ZEA resulted in elevated glutathione peroxidase (GPx) activity in the duodenal mucosa and kidney tissues, and an increased γ-glutamyltransferase (GGT) activity in the plasma, indicative of oxidative stress. In the liver tissue, no mycotoxin-induced response in GPx and thioredoxin reductase (TrxR) activities occurred, and the malondialdehyde (MDA) level was even reduced. Neither the plasma levels of retinol and α-tocopherol nor the activities of superoxide dismutase in erythrocytes and GPx in blood were affected in birds fed the contaminated diet. The only effect of lignin supplemented to the contaminated feed was that it prevented the increase of GPx activity in the duodenal mucosa as an indicator of oxidative stress.

Experimental addition of Eleutherococcus senticosus and probiotic to the canine diet

There is a current trend to support pet health through the addition of natural supplements to their diet, taking into account the high incidence of medical conditions related to their immune system and gastrointestinal tract. This study investigates effects of the plant Eleutherococcus senticosus as a dietary additive on faecal microbiota, faecal characteristics, blood serum biochemistry and selected parameters of cellular immunity in healthy dogs. A combination of the plant with the canine-derived probiotic strain Lactobacillus fermentum CCM 7421 was also evaluated. Thirty-two dogs were devided into 4 treatment groups; receiving no additive (control), dry root extract of E. senticosus (8 mg/kg of body weight), probiotic strain (108 CFU/mL, 0.1 mL/kg bw) and the combination of both additives. The trial lasted 49 days with 14 days supplementation period. Results confirm no antimicrobial effect of the plant on the probiotic abundance either in vitro (cultivation test) or in vivo. The numbers of clostridia, lactic acid bacteria and Gram-negative bacteria as well as the concentration of serum total protein, triglyceride, glucose and aspartate aminotransferase were significantly altered according to the treatment group. Leukocyte phagocytosis was significantly stimulated by the addition of probiotic while application of plant alone led to a significant decrease.

Identification and quantification of thymol metabolites in plasma, liver, and duodenal wall of broiler chickens using UHPLC‐ESI‐QTOF‐MS

In the present study, we aimed to develop a method for thymol sulfate and thymol glucuronide determination in plasma, liver and duodenal wall of broiler chickens after feeding with a Thymus vulgaris essential oil at the different concentrations (0.01, 0.05 and 0.1% w/w). UHPLC coupled with accurate-mass QTOF-MS was used for identification and quantification of thymol metabolites. Novel Waters Oasis Prime HLB solid-phase extraction cartridges were applied to sample clean-up with extraction recoveries ranged from 85 to 92%. The presence of thymol glucuronide was confirmed by MS software according to molecular formula, score, mass error and double bond equivalent. In terms of validation, calibration curves of thymol sulfate were constructed in matrix samples with linearity from 3.91 to 250.0 ng/mL and correlation coefficients were within the range of 0.9979-0.9995. Limits of detection were 0.97, 0.29 and 0.63 ng/mL and limits of quantification were 3.23, 0.97 and 2.09 ng/mL for plasma, liver and duodenal wall, respectively. Intra-day and inter-day precision expressed as relative standard deviation were <4.35%. To highlight, thymol metabolites were directly detected for the first time in liver and duodenal wall and this method was shown to be successfully applicable for investigation of thymol metabolism in chickens after thyme essential oil ingestion.