Gut-Faecal Microbial and Health-Marker Response to Dietary Fumonisins in Weaned Pigs.

This study investigated effects of dietary fumonisins (FBs) on gut and faecal microbiota of weaned pigs. In total, 18 7-week-old male pigs were fed either 0, 15 or 30 mg FBs (FB1 + FB2 + FB3)/kg diet for 21 days. The microbiota was analysed with amplicon sequencing of the 16S rRNA gene V3-V4 regions (Illumina MiSeq). Results showed no treatment effect (p > 0.05) on growth performance, serum reduced glutathione, glutathione peroxidase and malondialdehyde. FBs increased serum aspartate transaminase, gamma glutamyl-transferase and alkaline phosphatase activities. A 30 mg/kg FBs treatment shifted microbial population in the duodenum and ileum to lower levels (compared to control (p < 0.05)) of the families Campylobacteraceae and Clostridiaceae, respectively, as well as the genera Alloprevotella, Campylobacter and Lachnospiraceae Incertae Sedis (duodenum), Turicibacter (jejunum), and Clostridium sensu stricto 1 (ileum). Faecal microbiota had higher levels of the Erysipelotrichaceae and Ruminococcaceae families and Solobacterium, Faecalibacterium, Anaerofilum, Ruminococcus, Subdoligranulum, Pseudobutyrivibrio, Coprococcus and Roseburia genera in the 30 mg/kg FBs compared to control and/or to the 15 mg/kg FBs diets. Lactobacillus was more abundant in the duodenum compared to faeces in all treatment groups (p < 0.01). Overall, the 30 mg/kg FBs diet altered the pig gut microbiota without suppressing animal growth performance.

Micro-Encapsulated Microalgae Oil Supplementation Has No Systematic Effect on the Odor of Vanilla Shake-Test of an Electronic Nose.

In this study, we aimed to carry out the efficient fortification of vanilla milkshakes with micro-encapsulated microalgae oil (brand: S17-P100) without distorting the product's odor. A 10-step oil-enrichment protocol was developed using an inclusion rate of 0.2 to 2 w/w%. Fatty acid (FA) profile analysis was performed using methyl esters with the GC-MS technique, and the recovery of docosahexaenoic acid (C22:6 n3, DHA) was robust (r = 0.97, p < 0.001). The enrichment process increased the DHA level to 412 mg/100 g. Based on this finding, a flash-GC-based electronic nose (e-nose) was used to describe the product's odor. Applying principal component (PC) analysis to the acquired sensor data revealed that for the first four PCs, only PC3 (6.5%) showed a difference between the control and the supplemented products. However, no systematic pattern of odor profiles corresponding to the percentages of supplementation was observed within the PC planes. Similarly, when discriminant factor analysis (DFA) was applied, though a classification of the control and supplemented products, we obtained a validation score of 98%, and the classification pattern of the odor profiles did not follow a systematic format. Again, when a more targeted approach such as the partial least square regression (PLSR) was used on the most dominant sensors, a weak relationship (R2 = 0.50) was observed, indicating that there was no linear combination of the qualitative sensors' signals that could accurately describe the supplemented concentration variation. It can therefore be inferred that no detectable off-odor was present as a side effect of the increase in the oil concentration. Some volatile compounds of importance in regard to the odor, such as ethylacetate, ethyl-isobutarate, pentanal and pentyl butanoate, were found in the supplemented product. Although the presence of yeasts and molds was excluded from the product, ethanol was detected in all samples, but with an intensity that was insufficient to cause an off-odor.

A preliminary study on changes in heat shock protein 70 levels induced by Fusarium mycotoxins in rats: in vivo study.

The heat shock protein (Hsp70) level was assessed after 14 days of oral gavage-exposure to fumonisin B1 (FB1: 150 µg/animal/day), deoxynivalenol (DON: 30 µg/animal/day) and zearalenone (ZEN: 150 µg/animal/day), alone or in combinations (in additive manner: FD = FB1 + DON, FZ = FB1 + ZEN, DZ = DON + ZEN and FDZ = FB1 + DON + ZEN) in the liver, kidneys and lung of 24 adult male Wistar rats (n = 3/group). The liver was the most responsive tissue, as compared with kidney and lung. Except of DZ-treatment, mycotoxins elevated the Hsp70 levels in livers. The highest Hsp70-levels (≈ twofold) were in the DON, FD, FZ and FDZ treatments (additive effects). In the kidney, alterations (↑ ≈ twofold) were detected in ZEN, FD, FZ and DZ treatments. The least responsive organ was the lung (↑ only in FDZ, antagonistic effect). DON and ZEA exposures have altered the reduced glutathione concentration (↓) and glutathione peroxidase activity (↓) in the blood serum. The serum malondialdehyde level increased only after exposure to FD (synergistic effect), as compared with the DZ group (antagonistic effect). When the blood clinical chemistry was assessed, significant alterations were in alanine aminotransferase (80% increase in FDZ, antagonistic effect) and total protein (↓ ZEN). Results varied according to the organ, toxin type and interactions. Furthermore, oxidative stress was not the only key player behind the Hsp70 increase, in which another mechanism is suggested.

Assessment of Human Mycotoxin Exposure in Hungary by Urinary Biomarker Determination and the Uncertainties of the Exposure Calculation: A Case Study.

Urinary biomarkers of mycotoxin exposure were evaluated in the case of healthy people (n = 41) and coeliac patients (n = 19) by using a multi-biomarker LC-MS/MS immunoaffinity based method capable to analyse biomarkers of nine mycotoxins, i.e., fumonisin B1 (FB1), fumonisin B2 (FB2), deoxynivalenol (DON), zearalenone (ZEN), ochratoxin A (OTA), Aflatoxin B1 (AFB1), T-2 toxin, HT-2 toxin and Nivalenol (NIV). Urinary biomarker concentrations were used to calculate the probable daily intake (PDI) of fumonisin B1, deoxynivalenol, zearalenone and ochratoxin A and compared with their tolerable daily intake (TDI). The human urinary excretion rate values reported in the literature and the 24 h excretion rate measured in piglets were used to estimate and compare the PDI values of the four mycotoxins. The highest mean biomarker concentrations were found for DON (2.30 ng/mL for healthy people and 2.68 ng/mL for coeliac patients). Mean OTA concentration was significantly higher (p < 0.001) in healthy people compared to coeliac patients. PDI calculated with piglets excretion data exceeded the TDI values by a much smaller percentage than when they were calculated from human data, especially for FB1. The uncertainties arising from the different calculations can be well perceived on the basis of these data.

Interaction between the three frequently co-occurring Fusarium mycotoxins in rats.

To test the complex, acute biochemical effects of combined, naturally co-occurring fusariotoxins, a 5-day rat study was performed. Mycotoxin treatment was invented by intraperitoneal injection: FB1 (F): 9 µg/animal/day (approx. 30 µg/kg bw/day), DON (D): 16.5 µg/animal/day (approx. 55 µg/kg bw/day) and ZEN (Z): 12.75 µg/animal/day (approx. 42.5 µg/kg bw/day). The binary groups (FB1 and DON [FD], FB1 and ZEN [FZ] and DON and ZEN [DZ]) as well as the ternary (FB1 , DON and ZEN [FDZ]) group were dosed at the same combined level as the individual mycotoxins. Body weight, feed intake and mortality were not affected by any of the treatments. FB1 and DON in combination (FD) increased the plasma aspartate aminotransferase activity synergistically (compared to the individual FB1 and DON). In the liver, both the total glutathione (GSH) and the glutathione peroxidase (GPx) activity were increased (p < 0.05) by the binary FB1 and ZEN (FZ) and the DON and ZEN (DZ) groups as well as the ternary FB1 , DON and ZEA group (FDZ) compared to the control. The GSH level of the ternary group was significantly increased compared to the FB1 group, whereas the GPx activity of the ternary group was significantly increased compared to all three the individual mycotoxin groups. The Bliss independence method revealed synergism between DON and ZEN (DZ), as well as FB1 and DON (FD) on liver GPx activity. None of the toxins alone or in combination exerted strong genotoxicity on lymphocytes, neither on the gross histopathological characteristics. However, even at these low levels acute exposure of more than one of these mycotoxins (FB1 , DON and ZEN) affected metabolic and detoxification changes.

Fumonisin B1 exposure increases Hsp70 expression in the lung and kidney of rats without inducing significant oxidative stress.

The objective of this experiment was to determine whether fumonisin B1 (FB1) added to the diet of rats in a dose of 50 mg/kg changes the production of heat shock protein 70 (Hsp70) in the lungs and kidney of rats. We also studied the effect of this mycotoxin on the antioxidant system of the body. Mature (8 weeks old) male Wistar Crl:WI BR rats (n = 6/group) were fed the toxin-containing diet for 5 days. FB1 resulted in a 7% body weight reduction without significantly changing the feed intake. Western blot analysis of the lungs and kidney demonstrated a substantial (1.4-fold and 1.8-fold, respectively) increase in Hsp70 expression. Alterations could not be detected in the clinical chemical parameters (total protein, albumin, total cholesterol, glucose, creatinine and urea concentrations, and aspartate aminotransferase activity). There was no statistically significant change in malondialdehyde concentrations and the measured antioxidant parameters (the amount of reduced glutathione, GSH and glutathione peroxidase activity, GPx) in the blood plasma, lung and kidney tissue. Thus, it can be concluded that FB1 did not induce oxidative stress in the lungs and kidney, but increased Hsp70 production.

In vitro Interaction between Fumonisin B1 and the Intestinal Microflora of Pigs.

The caecal chyme of pigs was incubated anaerobically in McDougall buffer with and without fumonisin B1 (5 µg/ml) for 0, 24 and 48 h. The plate count agar technique was applied for enumerating the amount of bacteria including aerobic, anaerobic bacteria, coliform, Escherichia coli and Lactobacillus sp. The quantitative polymerase chain reaction was also performed to estimate the number of copies of the total bacteria, Lactobacillus, Bacteroides and Prevotella. No significant differences in the amount of bacterial groups between the experimental (buffer, chyme, and fumonisin B1) and control 1 groups (buffer + chyme) were observed in both methods. Fumonisin B1 and hydrolysed fumonisin B1 concentration were analysed by liquid chromatograghy - mass spectrometry. There was no significant difference in FB1 concentration between the experimental and the control 2 group (buffer and fumonisin B1) at 0 h incubation, 5.185 ± 0.174 µg/ml compared with 6.433 ± 0.076 µg/ml. Fumonisin B1 concentration in the experimental group was reduced to 4.080 ± 0.065 µg/ml at 24 h and to 2.747 ± 0.548 µg/ml at 48 h incubation and was significantly less than that of in the control group. Hydrolysed fumonisin B1 was detected after 24 h incubation (0.012 ± 0 µg/ml). At 48 h incubation time, hydrolysed fumonisin B1 concentration was doubled to 0.024 ± 0.004 µg/ml. These results indicate that fumonisin B1 can be metabolised by caecal microbiota in pigs though the number of studied bacteria did not change.

Physiological Effects of Whey- and Milk-Based Probiotic Yogurt in Rats.

In an in vitro experiment commercially available probiotic products were tested for the survival of bacteria under conditions of simulated human digestion either when used alone or mixed into yogurt. In the in vivo experiment the effects of feeding a whey- and milk-based yogurt prepared with the probiotic strain showing adequate survival in the in vitro experiment, was measured on body weight, feed con¬sumption and immune response of rats (IgG and IgA level after immunisation), on the composition and volatile fatty acid production of the intestinal microbiota and on the structure of intestinal villi. The Lactobacillus acidophilus (LA-15) strain had inadequate surviving ability in rats. Bifidobacterium animalis ssp. lactis (BB-12) improved the composition of the intestinal microflora, whereas whey-containing product had a mild immunostimulating effect and exerted a favourable influence on the morphology of intestinal villi. The consumption of yogurts increased the depth of crypts in the ileum, which resulted in enhanced secretion and thus softer faeces.

Effect of nursing methods and faeces consumption on the development of the bacteroides, lactobacillus and coliform flora in the caecum of the newborn rabbits.

The effect of nursing method and ingestion of maternal faeces on the development of the bacteroides, lactobacillus and coliform flora of the caecum in the first 10 days of life were examined in freely nursed pups having access to maternal faeces (Group FF), pups nursed once a day and having access (Group CF), or having no access (Group CN) to maternal faeces. Colonisation of the caecum by Bacteroides commenced already on day 3 after birth. On day 2 the bacteroides counts were below 100, while on day 4 they were already between 100 and 10,000. In Group CN, the Bacteroides counts were lower (by 14 to 40%) throughout the 10-day period studied than in the groups having access to maternal faeces. Differences between groups were significant only on days 4 and 6. The average number of maternal faecal pellets left behind the doe in Group CN was 3-4 (between 0.5 and 6.4 per doe). In Groups FF and CF the pellets became smaller, crumbled and finally disappeared from the nest box, they were consumed by the pups and could be found in their gastric content. The lactobacillus counts decreased in all three groups with age, from 6.0 to 3.5 log10 CFU.g-1 (FF), 4.6 to 2.8 log10 CFU.g-1 (CF) and 5.1 to 3.1 log10 CFU.g-1 (CN), respectively. The coliform counts were higher in the first 4 days in FF (5.6 log10 CFU.g-1) than in CF (< 2 log10 CFU.g-1) and CN (2-3.6 log10 CFU.g-1) animals. Bacteroides could be cultured from the surface of the vulvar labia (max. 1000 colony count) and the vagina (max. 190 colony count), so young rabbits could become "infected" by them already in the doe's vagina. Thus prevention of ingestion of maternal faeces only slightly influenced the development of the bacteroides flora, the faeces left behind by the doe did not play an exclusive role in their colonisation.