In Vitro Evaluation of Phytobiotic Mixture Antibacterial Potential against Enterococcus spp. Strains Isolated from Broiler Chicken.

Enterococcus spp. are normal intestinal tract microflorae found in poultry. However, the last decades have shown that several species, e.g., Enterococcus cecorum, have become emerging pathogens in broilers and may cause numerous losses in flocks. In this study, two combinations (H1 and H2) of menthol, 1,8-cineol, linalool, methyl salicylate, γ-terpinene, p-cymene, trans-anethole, terpinen-4-ol and thymol were used in an in vitro model, analyzing its effectiveness against the strains E. cecorum, E. faecalis, E. faecium, E. hirae and E. gallinarum isolated from broiler chickens from industrial farms. To identify the isolated strains classical microbiological methods and VITEK 2 GP cards were used. Moreover for E. cecorum a PCR test was used.. Antibiotic sensitivity (MIC) tests were performed for all the strains. For the composition H1, the effective dilution for E. cecorum and E. hirae strains was 1:512, and for E. faecalis, E. faecium and E. gallinarum, 1:1024. The second mixture (H2) showed very similar results with an effectiveness at 1:512 for E. cecorum and E. hirae and 1:1024 for E. faecalis, E. faecium and E. gallinarum. The presented results suggest that the proposed composition is effective against selected strains of Enterococcus in an in vitro model, and its effect is comparable to classical antibiotics used to treat this pathogen in poultry. This may suggest that this product may also be effective in vivo and provide effective support in the management of enterococcosis in broiler chickens.

The Impact of a Phytobiotic Mixture on Broiler Chicken Health and Meat Safety.

The purpose of the study was to assess the effects of different doses of a phytobiotic mixture on selected production parameters and meat quality and to assess the residue of the preparation in tissues and the possible toxic effects in broiler chickens. Broiler chicks aged 160 days, divided into four equal groups, were supplemented with the phytobiotic mixture at different doses, D1-0.5 mL/L, D2-1 mL/L, and D3-2 mL/L, four times during a 42-day trial. There were no statistically significant differences in weight gain per week of life and mortality in the birds. The study also demonstrated that the use of the mixture of phytobiotics had no significant effect on colour, pH, WHC, and natural leakage. However, a beneficial effect of the additive was found in the group treated with a dose of 1 mL/L, where less thermal leakage from the meat was demonstrated. Furthermore, significant differences in the change in thigh muscle tenderness were also observed. In the histopathological analysis of the liver no significant differences were observed. In addition, no residues of the mixture or its metabolites were found in the tissues analysed. In conclusion, the proposed scheme of administration of the phytobiotic additive, regardless of the dose, does not cause pathological changes in organs and does not carry the risk of residues of the product in tissues intended for human consumption.

In Vitro Evaluation of Antiprotozoal Properties, Cytotoxicity Effect and Anticancer Activity of New Essential-Oil Based Phytoncide Mixtures.

Protozoa, in both humans and animals, are one of the leading causes of disease. International programmes introduced in many countries have helped reduce the incidence of disease. However, it has recently become increasingly difficult to achieve the goals set for the coming years. One of the main reasons for this, as with other pathogenic organisms, such as bacteria and fungi, is the increasing resistance to current methods of treating and preventing infection. Therefore, new therapies with high efficacy are needed. In the present study, the novel mixtures of essential oils (EOs), clove, garlic, Ceylon cinnamon, and rosemary with organic acids (acetic, propionic, lactic) and metal ions (Cu, Mn, Zn) were tested against five selected model protozoa (Euglena gracilis, Gregarina blattarum, Amoeba proteus, Paramecium caudatum, Pentatrichomonas hominis). The cytotoxicity and potential anticancer activity of the obtained combinations were tested on the human fibroblasts (NHDF) and human cancer cell lines (A549, MCF7, LoVo, HT29). All of the mixtures showed very good antiprotozoal properties. The most efficient were the combination of clove and rosemary essential oils, mixtures of acids, and Mn ions. The LD50 values were in the range of 0.001-0.006% and the LD100 values were 0.002-0.008%. All of the tested mixtures did not show cytotoxicity against normal cells, but did show growth inhibition against cancer cell lines. The most cytotoxic against cancer cells were combinations with cinnamon essential oil. Nevertheless, the proposed combinations containing essential oils, organic acids, and metal ions have high antiprotozoal activity, with low toxicity to healthy human cells.

In Vitro Assessment of Antimicrobial Activity of Phytobiotics Composition towards of Avian Pathogenic Escherichia coli (APEC) and Other E. coli Strains Isolated from Broiler Chickens.

Escherichia coli infections (including APEC) in broiler chickens are not only a health and economic problem of the flock, but also a significant health threat to poultry meat consumers. The prophylactic and therapeutic effects of the phytobiotic composition on E. coli in broiler chickens were previously described. However, most of the data were related to the reference strains (for both in vitro and in vivo models). Based on the previous studies in human and animals, E. coli strains seem to be multidrug resistance. This, in turn, makes it necessary to develop effective alternative methods of treating this type of infection already at the stage of poultry production. In the present study, the antibacterial activity against various strains of E. coli (including APEC) was assessed for two innovative phytobiotics mixtures: H1, containing thymol, menthol, linalool, trans-anethole, methyl salicylate, 1,8-cineol, and p-cymene; H2, in addition to compounds from H1, containing terpinen-4-ol and γ-terpinene. The unique mixtures of phytobiotics used in the experiment were effective against various strains of E. coli, also against APEC, isolated from broiler chickens from traditional industrial breeding, as well as against those showing colistin resistance. The minimum inhibitory concentration (MIC) values for these unique mixtures were: For H1 1:512 for APEC and non-APEC E. coli strains isolated from day old chicks (DOCs), 1:512 for non-APEC, and 1:1024 for non-APEC isolated from broilers sample. For mixture H2, MIC for APEC from both type of samples (DOCs and broilers) was 1:1024 and for non-APEC (DOCs and broilers) was 1:512. The results suggest that phytobiotic compositions used in this study can be successfully used as a natural alternative to antibiotics in the treatment of E. coli infections in broiler chickens. The promising results may be a crucial point for further analyses in broiler flocks exposed to E. coli infections and where it is necessary to reduce the level of antibiotics or completely eliminate them, thus reducing the risk of foodborne infections.

In Vitro Evaluation of Antimicrobial Effect of Phytobiotics Mixture on Salmonella spp. Isolated from Chicken Broiler.

BACKGROUND: The identification of natural antibacterial agents from various sources that can act effectively against disease-causing foodborne bacteria is one of the major concerns throughout the world. In the present study, a unique phytobiotics mixture containing thymol, menthol, linalool, trans-anethole, methyl salicylate, 1,8-cineole, and p-cymene was evaluated for antibacterial activity against selected strains of Salmonella spp. RESULTS: The phytobiotics mixture was effective against Salmonella enterica subsp. enterica serovars Enteritidis, Typhimurium, and Kentucky. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of this unique mixture for these three pathogens were 1:256. Among these three strains, one S. Kentucky presented the most extensive resistance profiles to 18 antibiotics belonging to 5 classes of antibiotics. One of S. Typhimurium presents extensive resistance profiles to 14 antibiotics belonging to 5 classes of antibiotics. CONCLUSIONS: The results suggest that the phytobiotics mixture used in the experiment can be used as a strong natural antibacterial agent against Gram-negative foodborne pathogens such as S. Typhimurium, S. Kentucky, and S. Enteritidis. This is a preliminary analysis of the effectiveness of a phytobiotic product in an in vitro model, which may be the starting point for further studies, including in vivo animal models.

Novel Formula of Antiprotozoal Mixtures.

Antimicrobial resistance (AMR) is becoming more common in both bacteria and pathogenic protozoa. Therefore, new solutions are being sought as alternatives to currently used agents. There are many new ideas and solutions, especially compounds of natural origin, including essential oils. In the present study, the antiprotozoal activity of a mixture of essential oils (eucalyptus, lavender, cedar and tea tree), organic acids (acetic acid, propionic acid and lactic acid) and metal ions (Cu, Zn, Mn) were tested. As a model, protozoans were selected: Euglena gracilis, Gregarina blattarum, Amoeba proteus, Paramecium caudatum, Pentatrichomonas hominis. The tested concentrations of mixtures were in the range of 0.001-1.5%. The analyses show unexpected, very strong protozoicidal activity of combinations, presenting the synergy of compounds via determination of LD50 and LD100 values. Obtained mixtures showed significantly higher activity against protozoans, compared to chloramphenicol and metronidazole. Most of the analyzed samples show high antiprotozoal activity at very low concentration, in the range of 0.001-0.009%. The most effective combinations for all analyzed protozoans were the cedar essential oil and tea tree essential oil with a mixture of acids and manganese or zinc ions. Innovative combinations of essential oils, organic acids and metal ions are characterized by very high antiprotozoal activity at low doses, which, after further investigation, can be applicable for control of protozoan pathogens.

The Use of Selenium Yeast and Phytobiotic in Improving the Quality of Broiler Chicken Meat.

The aim of the present study was to investigate the effect of selenium yeast and phytobiotic on the storage capacity, selected quality parameters of meat and content of selenium in muscles obtained from broilers. In the experiment, 1440 male broiler chickens (Ross 308) were randomly assigned to four research groups: group received no additive (G1), group received a supplement of 0.3 mg Se (as sodium selenite)/kg of feed mixture (G2), group received 0.2 g phytobiotic and 0.3 mg Se as 0.1 g selenium yeast per 1 kg of feed mixture (G3) and group received 0.3 mg Se as 0.1 g selenium yeast per 1 kg of feed mixture (G4). Measurement of pH, determination of water retention capacity, degree of advancement of oxidative changes and selenium content in muscles were performed. Samples of chickens' breast and thigh muscles were microbiologically analyzed. The additives significantly influenced the level of oxidation in muscles and the incorporation of selenium. The meat of chickens receiving organic selenium was characterized by significantly lower dynamics of oxidative changes. The studies carried out showed that selenium in organic form had better absorption.

Volatile Composition and Sensory Properties as Quality Attributes of Fresh and Dried Hemp Flowers (Cannabis sativa L.).

Flowers of hemp (Cannabis sativa L.) are widely used in cosmetics, food, and in the pharmaceutical industry. The drying process plays a key role in retention of aroma and also in the quality of products. Seven variants of hemp flower drying, including convection drying (CD), vacuum-microwave drying (VMD), and combined drying consisting of convective pre-drying followed by vacuum-microwave finishing drying (CPD-VMFD) were checked in this study. For each process, we applied the two-term model. Dried material was submitted to color and chromatographical assessments. Analyses of obtained essential oil showed the presence of 93 volatile compounds, predominantly ß-myrcene, limonene, and ß-(E)-caryophyllene, as well as α-humulene. Application of 240 W during VMD and 50 °C during CD gave the highest retention of aroma compounds, amounting to 85 and 76%, respectively, but with huge color changes. Additionally, sensory analysis proved that drying with a microwave power of 240 W provides a product most similar to fresh material.

Chemical profiling and cytotoxic activity of 150-year old original sample of Jerusalem Balsam.

Herbal formulations have been used in ethnomedicine and pharmacy around the world for thousands of years. One of them is Jerusalem Balsam (JB), which came into use in the seventeenth century. Today, people still produce and use it regularly as prophylactic supplement. JB has been widely used in Europe since the nineteenth century, as a remedy possessing antibacterial, antifungal and anti-inflammatory activities. The composition of the product was not known, although possible formulations were reported. In this study the original sample, which dated back to 1870, was investigated for chemical composition and cytotoxic activity. The obtained results were compared with results from more recently produced samples. Several tests were carried out, namely GC-MS, UPLC-PDA-Q-TOF-MS and MTT. Only the 150-year old sample showed a significant cytotoxic activity on cancer cell lines. At a concentration of 125 µg/mL after 72 h of incubation, the original sample inhibited almost 90% of cell metabolic activity, while contemporary samples showed none or little activity. None of the tested samples showed a significant impact on normal cells. These results may be attributed to the activities of benzoic acid and its derivatives, cinnamic acid derivatives, vanillin, group of sesquiterpenes and cembrene.