Comparison of the Effects of Essential Oils from Cannabis sativa and Cannabis indica on Selected Bacteria, Rumen Fermentation, and Methane Production-In Vitro Study.

This study aimed to evaluate the effects of essential oils (EOs) extracted from Cannabis sativa L. and Cannabis indica Lam. on in vitro ruminal fermentation characteristics, selected rumen microbial populations, and methane production. GC-MS analyses allowed us to identify 89 compounds in both EOs. It was found that E-ß-caryophyllene predominated in C. sativa (18.4%) and C. indica (24.1%). An in vitro (Ankom) test was performed to analyse the control and monensin groups, as well as the 50 µL or 100 µL EOs. The samples for volatile fatty acids (VFAs), lactate, and microbiological analysis were taken before incubation and after 6 and 24 h. The application of EOs of C. indica resulted in an increase in the total VFAs of acetate and propionate after 6 h of incubation. The applied EOs had a greater impact on the reduction in methane production after 6 h, but no apparent effect was noted after 24 h. Lower concentrations of C. sativa and C. indica had a more pronounced effect on Lactobacillus spp. and Buryrivibrio spp. than monensin. The presented findings suggest that C. sativa and C. indica supplementation can modify ruminal fermentation, the concentrations of specific volatile fatty acids, and methane production.

spa Types and Staphylococcal Enterotoxin Production of Staphylococcus aureus Isolated from Wild Boar.

Little is known about the structure of S. aureus population and the enterotoxin gene content in wild boar. In 1025 nasal swabs from wild boars, 121 S. aureus isolates were identified. Staphylococcal enterotoxin (SE) genes were identified in 18 isolates (14.9%). The seb gene was found in 2 S. aureus isolates, sec in 2 isolates, the see and seh genes were found in 4 and 11 isolates, respectively. The production of SEs was evaluated in bacteria grown in microbial broth. Concentration of SEB reached 2.70 µg/ml after 24 h and 4.46 µg/ml at 48 h. SEC was produced at 952.6 ng/ml after 24 h and 7.2 µg/ml at 48 h. SEE reached 124.1 ng/ml after 24 h and 191.6 ng/ml at 48 h of culture. SEH production reached 4.36 µg/ml at 24 h and 5.42 µg/ml at 48 h of culture. Thirty-nine spa types were identified among S. aureus isolates. The most prevalent spa types were t091 and t1181, followed by t4735 and t742, t3380 and t127. Twelve new spa types, i.e., t20572‒t20583 were identified. The wild boar S. aureus population was shown to contain previously identified animal/human-associated spa types and spa types not identified in humans or animals. We also indicate that wildlife animals can be a significant reservoir of see-positive S. aureus.

Stability and Resistance to Proteolysis of Enterotoxins SEC and SEL Produced by Staphylococcus epidermidis and Staphylococcus aureus.

The only staphylococcal enterotoxins produced by Staphylococcus epidermidis include SECepi and SELepi, whereas Staphylococcus aureus produces orthologous SECs and SEL having different sequences. We compared S. epidermidis and S. aureus SECs and SELs in terms of resistance to proteolysis and both, thermal and chemical stability. We show that SECepi and SELepi produced by S. epidermidis have similar resistance to proteolysis if compared with their respective orthologues produced by S. aureus. Studied S. epidermidis and S. aureus SEC variants incubated with pepsin at pH 2.0 were found to be more resistant to proteolysis than SELs. SELs turned out to be more resistant than SECs to proteolysis with trypsin at pH 8.0. SECepi was found to be more resistant to thermal denaturation if compared with its S. aureus orthologues. The S. epidermidis and S. aureus SEC variants were found to have higher thermal stability than SELs. Our data indicate that, due to their high stability, the enterotoxins SECepi and SELepi produced in food by S. epidermidis may pose a food safety risk comparable with that posed by S. aureus enterotoxins.

Staphylococcal Enterotoxin Genes in Coagulase-Negative Staphylococci-Stability, Expression, and Genomic Context.

In the current study, we screened a collection of coagulase-negative staphylococci (CoNS) isolates for orthologues of staphylococcal enterotoxins (SEs) involved in S. aureus-related staphylococcal food poisoning (SFP). The amplicons corresponding to SEs were detected in S. chromogenes, S. epidermidis, S. haemolyticus, S. borealis, S. pasteuri, S. saprophyticus, S. vitulinus, S. warneri, and S. xylosus. All amplicons were sequenced and identified as parts of known S. aureus or S. epidermidis SE genes. Quantitative real-time PCR allowed determining the relative copy number of each SE amplicon. A significant portion of the amplicons of the sea, seb, sec, and seh genes occurred at low copy numbers. Only the amplicons of the sec gene identified in three isolates of S. epidermidis displayed relative copy numbers comparable to sec in the reference enterotoxigenic S. aureus and S. epidermidis strains. Consecutive passages in microbiological media of selected CoNS isolates carrying low copy numbers of sea, seb, sec, and seh genes resulted in a decrease of gene copy number. S. epidermidis isolates harbored a high copy number of sec, which remained stable over the passages. We demonstrated that enterotoxin genes may occur at highly variable copy numbers in CoNS. However, we could identify enterotoxin genes only in whole-genome sequences of CoNS carrying them in a stable form at high copy numbers. Only those enterotoxins were expressed at the protein level. Our results indicate that PCR-based detection of enterotoxin genes in CoNS should always require an additional control, like analysis of their presence in the bacterial genome. We also demonstrate S. epidermidis as a CoNS species harboring SE genes in a stable form at a specific chromosome site and expressing them as a protein.

Regulatory and Enterotoxin Gene Expression and Enterotoxins Production in Staphylococcus aureus FRI913 Cultures Exposed to a Rotating Magnetic Field and trans-Anethole.

The study aimed to examine the influence of a rotating magnetic field (RMF) of two different frequencies (5 and 50 Hz) on the expression of regulatory (agrA, hld, rot) and staphylococcal enterotoxin (SE-sea, sec, sel) genes as well as the production of SEs (SEA, SEC, SEL) by the Staphylococcus aureus FRI913 strain cultured on a medium supplemented with a subinhibitory concentration of trans-anethole (TA). Furthermore, a theoretical model of interactions between the bacterial medium and bacterial cells exposed to RMF was proposed. Gene expression and SEs production were measured using quantitative real-time PCR and ELISA techniques, respectively. Based on the obtained results, it was found that there were no significant differences in the expression of regulatory and SE genes in bacteria simultaneously cultured on a medium supplemented with TA and exposed to RMF at the same time in comparison to the control (unexposed to TA and RMF). In contrast, when the bacteria were cultured on a medium supplemented with TA but were not exposed to RMF or when they were exposed to RMF of 50 Hz (but not to TA), a significant increase in agrA and sea transcripts as compared to the unexposed control was found. Moreover, the decreased level of sec transcripts in bacteria cultured without TA but exposed to RMF of 50 Hz was also revealed. In turn, a significant increase in SEA and decrease in SEC and SEL production was observed in bacteria cultured on a medium supplemented with TA and simultaneously exposed to RMFs. It can be concluded, that depending on SE and regulatory genes expression as well as production of SEs, the effect exerted by the RMF and TA may be positive (i.e., manifests as the increase in SEs and/or regulatory gene expression of SEs production) or negative (i.e., manifests as the reduction in both aforementioned features) or none.

Hemosiderin Accumulation in Liver Decreases Iron Availability in Tachycardia-Induced Porcine Congestive Heart Failure Model.

Despite advances in the management of iron deficiency in heart failure (HF), the mechanisms underlying the effects of treatment remain to be established. Iron distribution and metabolism in HF pathogenesis need to be clarified. We used a porcine tachycardia-induced cardiomyopathy model to find out how HF development influences hepatic and myocardial iron storing, focusing on ferritin, the main iron storage protein. We found that cumulative liver congestion (due to the decrease of heart function) overwhelms its capacity to recycle iron from erythrocytes. As a consequence, iron is trapped in the liver as poorly mobilized hemosiderin. What is more, the ferritin-bound Fe3+ (reflecting bioavailable iron stores), and assembled ferritin (reflecting ability to store iron) are decreased in HF progression in the liver. We demonstrate that while HF pigs show iron deficiency indices, erythropoiesis is enhanced. Renin-angiotensin-aldosterone system activation and hepatic hepcidin suppression might indicate stress erythropoiesisinduced in HF. Furthermore, assembled ferritin increases but ferritin-bound Fe3+ is reduced in myocardium, indicating that a failing heart increases the iron storage reserve but iron deficiency leads to a drop in myocardial iron stores. Together, HF in pigs leads to down-regulated iron bioavailability and reduced hepatic iron storage making iron unavailable for systemic/cardiac needs.

Antioxidant defence and oxidative stress markers in cats with asymptomatic and symptomatic hypertrophic cardiomyopathy: a pilot study.

BACKGROUND: Hypertrophic cardiomyopathy is the most common cardiovascular cause of death in cats. Although the majority of cats remain asymptomatic, some may develop signs of chronic heart failure due to diastolic failure, arterial thromboembolism (ATE) or sudden cardiac death. Therefore, it is crucial to identify individuals that are in high risk of developing cardiac complications before the onset of life-threatening signs. Oxidative stress is the imbalance between the production and neutralisation of reactive oxygen species. Uncontrolled reactive oxygen species overproduction leads to protein and lipid peroxidation and damages the DNA strands, injuring the cells and leading to their death. The aim of the study was to evaluate the oxidative state in cats with hypertrophic cardiomyopathy and healthy controls. RESULTS: In total, 30 cats divided into three groups were assessed: animals with clinically evident hypertrophic cardiomyopathy (HCM; n = 8), subclinical hypertrophic cardiomyopathy (SUB-HCM; n = 11) and healthy controls (n = 11). The activity of superoxide dismutase was statistically significantly lower in animals with symptomatic and asymptomatic hypertrophic cardiomyopathy (HCM 0.99 ± 0.35 U/mL; SUB-HCM 1.39 ± 0.4 U/mL) compared to healthy cats (2.07 ± 0.76 U/mL, p < 0.01). The activity of catalase was significantly lower in the SUB-HCM group (19.4 ± 4.2 nmol/min/mL) compared to the HCM (23.6 ± 5.9 nmol/min/mL) and the control (30 ± 7.5 nmol/min/mL, p < 0.01) group. The activity of glutathione peroxidase was 4196 ± 353 nmol/min/mL in the HCM group, 4331 ± 451 nmol/min/mL in the SUB-HCM group and 4037 ± 341 nmol/min/mL in the control group and did not differ significantly between groups. The total antioxidant capacity of plasma was 602 ± 65.5 copper reducing equivalents (CRE) in the HCM group, 605.9 ± 39.9 CRE in the SUB-HCM group and 629 ± 77.5 CRE in the healthy cats and did not differ significantly between the groups. CONCLUSIONS: Activities of superoxide dismutase and catalase differed in cats with hypertrophic cardiomyopathy, however the activity of the latter was only significantly lower in asymptomatic stage of the disease. The potentially beneficial effect of antioxidative substances on the disease progression in the asymptomatic and symptomatic stage of this disease should also be examined.

Do You Know What You Eat? Kebab Adulteration in Poland.

In recent years, consumer interest in meat authenticity has increased. Fraudulent claims are most likely to be regarding meat origin, meat substitution, meat processing treatment, and non-meat ingredient additions. This study focuses on the substitution of meat species in processed kebab-like food sales in Poland. The growing popularity of kebab-like foods and the limited number of official inspections of this type of food make this topic interesting. In this study, the results reveal that 60% of the foods analyzed contain an undeclared ingredient or the substitution of an expensive ingredient with a cheaper option.

Enhancement of neutrophil chemotaxis by trans-anethole-treated Staphylococcus aureus strains.

This study aimed to analyze the chemotactic response of differentiated HL-60 neutrophil-like (dHL-60) cells to trans-anethole (TA)-treated Staphylococcus aureus strains. Special attention was paid to evaluate the influence of TA on the chp gene expression level, as well as molecular docking and molecular dynamics (MD) simulation studies on interactions of TA with chemotaxis inhibitory protein of S. aureus (CHIPS). The following parameters were studied: susceptibility to TA using the agar diffusion method, the chp gene detection and its expression under TA influence, and clonal diversity of S. aureus strains using molecular techniques. Furthermore, a chemotactic response of dHL-60 cells to TA-treated S. aureus using Boyden chamber assay was detected and molecular modeling using both the docking methodology and unbiased MD simulations was conducted. It was found that TA showed antibacterial activity against all strains. Three genotypes and one unique pattern were distinguished among the strains. 50% of the isolates were chp-positive. It was observed that TA reduced/inhibited chp gene expression in most S. aureus strains. Enhanced chemotactic response of dHL-60 cells to TA-treated S. aureus strains was also noted. This correlation was similar for both chp-positive and chp-negative strains. Both molecular docking and MD simulations studies confirmed that TA is preferentially bound in the complement component 5a/CHIPS interface interaction region and can interfere with any processes exploiting this binding cavity. It has been proven that dHL-60 cells exhibited a higher chemotactic response to TA-treated S. aureus strains in comparison to non-treated bacteria, regardless of the achieved expression of the chp gene or its lack. Nevertheless, further analyses are required to understand this mechanism better.

Effect of low-dose hydrocortisone and inhaled nitric oxide on inflammatory mediators and local pulmonary metalloproteinases activity in LPS-induced sepsis in piglets.

Hospital mortality in sepsis varies between 30-45%. It has been shown that administration of inhaled nitric oxide (iNO) and intravenous corticosteroid in a porcine endotoxemia model attenuated the systemic inflammatory response. We explored the anti-inflammatory effect of a double-treatment strategy (iNO + low-dose steroid) on the lungs in a long-term porcine endotoxic shock model. As metalloproteinases (MMPs) are involved in the initiation of multiple organ dysfunction in septic shock, we evaluated the influence of this combination therapy on MMP2 and MMP9 activity and proIL-1ß maturation. A shock-like condition was established in 23 animals by continuous infusion of E. coli lipopolysaccharide (LPS) for 10 h. Then the animals were observed for 10 h. Twelve pigs received iNO and hydrocortisone (iNO treatment started 3 h after the initial LPS infusion and continued until the end of the experiment). Eleven pigs were controls. Pigs treated with iNO and hydrocortisone displayed less inflammatory infiltrates in the lungs than the controls and a lower level of IL-1ß. The proMMP2 was significantly decreased in the iNO and hydrocortisone group. The amount of an active MMP9 (~ 60 kDa) was decreased in the iNO and hydrocortisone group. Total gelatinolytic activity was lower in the iNO and hydrocortisone group. Reduced MMP activity was accompanied by a 2.5-fold decrease of the active IL-1ß form (17 kDa) in the pulmonary tissue of iNO combined with hydrocortisone exposed pigs. We demonstrated that in a porcine endotoxemia model the NO inhalation combined with intravenous hydrocortisone led to the attenuation of the inflammatory cascade induced by bacterial LPS. The decrease in pulmonary MMPs activities was accompanied by reduced proIL-1ß processing.

Analysis of enterotoxigenic effect of Staphylococcus aureus and Staphylococcus epidermidis enterotoxins C and L on mice.

Pathogenic properties of orthologues to S. aureus staphylococcal enterotoxin C (SEC) and staphylococcal enterotoxin L (SEL) produced by S. epidermidis are largely unexplored. We assessed the enteropathogenic effects of S. epidermidis SECepi and SELepi and S. aureus SEC3 and SEL after oral administration to Balb/c mice. Intestinal sections from SE-treated mice were analyzed histopathologically. The T cell lineage markers (αß and γδ TCR CD3, CD4, CD8), T-cell activation marker CD69 and proliferation-related marker CD71 were assessed in intraepithelial lymphocytes (IEL), mesenteric lymph nodes (MLN) and spleens (SPL). Serum concentrations of SEC and SEL were determined. Ortologous S. epidermidis and S. aureus SEs exerted a number of common histopathological changes in the mouse gut. Atrophy, generation of villi gap and edema of the villi were the most prominent effects of SE treatment observed in mouse gut sections. The most marked effect of ortologous S. epidermidis and S. aureus SEs on the number of goblet cells, crypt depth and villi height was noted in the mice duodenum and jejunum. We indicate early changes of TCRαß CD4-CD8a+ T and TCRαß CD4+CD8a+ T cells in response to both S. aureus and S. epidermidis SEs. Upon the treatment with SEs, markers of T cell activation and proliferation were upregulated in both αß and γδ T cell populations derived from IEL and MLN. We demonstrated that S. epidermidis-encoded SEs applied via oral route exert pathological changes in mice gut similarly to S. aureus-encoded SEs. For the first time we indicated that SEL co-produced together with SEC by both S. aureus and S. epidermidis induces some elements of mice gut immune response and contributes to gastrointestinal tract damage. Our results indicate the potential involvement of CoNS-encoded enterotoxins in the pathogenesis of SFP.

Serum Total Antioxidant Capacity and Enzymatic Defence of Dogs with Chronic Heart Failure and Atrial Fibrillation: A Preliminary Study.

INTRODUCTION: Atrial fibrillation may potentially contribute to oxidative stress to a greater extent than chronic heart failure. The aim of the study was to compare the serum total antioxidant capacity and enzymatic antioxidant defence of dogs with chronic heart failure and atrial fibrillation with those of subjects with chronic heart failure and sinus rhythm and healthy controls. MATERIAL AND METHODS: A total of 33 dogs were divided into three groups: dogs with chronic heart failure and atrial fibrillation (CHF + AF; n = 12), chronic heart failure and sinus rhythm (CHF + SR; n = 9), and healthy controls (n = 12). Serum total antioxidant capacity (TAC), serum CuZn-superoxide dismutase (SOD) and catalase, and plasma glutathione peroxidase (GPx) activity were determined. RESULTS: SOD activity and serum TAC were significantly lower in the study groups than in control animals. Catalase activity was significantly higher and plasma GPx activity significantly lower in dogs with CHF + AF compared with the CHF + SR and control dogs. CONCLUSION: The results suggest that chronic heart failure in dogs significantly impacts the serum TAC and the antioxidant enzymatic defence, while plasma GPx activity is markedly lower in dogs with chronic heart failure and atrial fibrillation. The role of that imbalance needs further investigation.

Antioxidative enzyme activity and total antioxidant capacity in serum of dogs with degenerative mitral valve disease.

This study was designed to evaluate the antioxidative status of serum by measuring its total antioxidant capacity, as well as the antioxidant enzyme activity (superoxide dismutase, catalase, and glutathione reductase), in dogs with various stages of degenerative mitral valve disease (DMVD) compared to healthy controls. In total, 71 client-owned dogs in different stages of DMVD, which included healthy controls, took part in the study. Following an anamnesis, clinical examination, standard transthoracic echocardiograpic examination, chest X-ray, complete blood (cell) count, and serum biochemistry, dogs were divided into 2 study groups. Blood was drawn from each dog once at the time of presentation and selected antioxidant parameters were measured using commercially available assay kits. The activity of superoxide dismutase gradually decreased in the more advanced stages of DMVD, while the activity of catalase was significantly higher in the group of dogs with asymptomatic DMVD compared to healthy controls and dogs with symptomatic DMVD. No significant changes were noted in total antioxidant capacity and the activity of glutathione reductase. Results suggested that DMVD has a significant impact on the activity of superoxide dismutase and catalase in the serum of the tested dogs. Knowledge of changes in the activity of antioxidative enzymes may warrant further studies, possibly to evaluate the potential role of compounds with antioxidative properties in the clinical outcome of dogs with DMVD.

La présente étude a été conçue afin d'évaluer le statut antioxydant du sérum en mesurant sa capacité antioxydante totale, ainsi que l'activité antioxydante enzymatique (superoxyde dismutase, catalase, et glutathion réductase), chez des chiens avec des degrés divers de maladie dégénérative de la valvule mitrale (DMVD) comparativement à des témoins en santé. Au total, 71 chiens appartenant à des clients à différents stades de DMVD, qui incluaient des témoins en santé, ont pris part à cette étude. À la suite de la prise d'anamnèse, d'un examen clinique, d'un examen échocardiographie transthoracique standard, de radiographie thoracique, d'un comptage cellulaire sanguin complet, et d'analyse biochimique sérique, les chiens étaient séparés en deux groupes d'étude. Du sang fut prélevé de chaque chien une fois au moment de la présentation et les paramètres antioxydants sélectionnés furent mesurés à l'aide d'une trousse disponible commercialement. L'activité de la superoxyde dismutase diminuait graduellement dans les stades plus avancés de DMVD, alors que l'activité de la catalase était significativement plus élevée dans le groupe de chiens avec une DMVD asymptomatique comparativement aux témoins en santé et aux chiens avec une DMVD symptomatique. Aucun changement significatif n'était noté dans la capacité antioxydante totale et dans l'activité de la glutathion réductase. Les résultats suggèrent que la DMVD a un impact significatif sur l'activité de la superoxyde dismutase, et de la catalase dans le sérum des chiens testés. Des connaissances sur les changements dans l'activité des enzymes antioxydantes pourraient justifier des études additionnelles, possiblement pour évaluer le rôle potentiel de produits avec des propriétés antioxydantes dans le devenir clinique de chiens avec DMVD.(Traduit par Docteur Serge Messier).

Antimicrobial Resistance and Biofilm Formation Capacity of Salmonella enterica Serovar Enteritidis Strains Isolated from Poultry and Humans in Poland.

Salmonella enterica ser. Enteritidis (S. enterica ser. Enteritidis) is the most frequently detected serovar in human salmonellosis, and its ability to produce a biofilm and the risk of transmission from animals and food of animal origin to humans are significant. The main aim of the present work was to compare S. enterica ser. Enteritidis strains isolated from poultry and human feces in terms of resistance profiles, prevalence of selected resistance genes, and their potential for biofilm formation, by assessing their biofilm growth intensity, the prevalence and expression of selected genes associated with this phenomenon, and the correlation between increased antimicrobial resistance and biofilm formation ability of the two tested groups of S. enterica ser. Enteritidis. This study showed a difference in antimicrobial resistance (minimal inhibitory concentration value) between S. enterica ser. Enteritidis groups; however, the majority of multidrug-resistant (MDR) strains were isolated from poultry (environmental samples from chicken broilers, turkey broilers, and laying hens). Differences in the prevalence of resistance genes were observed; the most common gene among poultry strains was floR, and that among strains from humans was blaTEM. S. enterica ser. Enteritidis strains isolated from poultry under the tested incubation conditions exhibited better biofilm growth than strains isolated from humans. A higher level of gene expression associated with the production of cellulose was only detected in the S48 strain isolated from poultry. On the other hand, increased expression of genes associated with quorum sensing was observed in two strains isolated from poultry farms and one strain isolated from human feces.