Niya Zhang

In vitro investigation of individual and combined cytotoxic effects of aflatoxin B1 and other selected mycotoxins on the cell line porcine kidney 15.

In the present study, we evaluated the nephrotoxicity of individual mycotoxins and combinations of aflatoxin B1 (AFB1), zearalenone (ZEA), deoxynivalenol (DON), and fumonisin B1 (FB1) to livestock using porcine kidney 15 cells (PK-15) as a disease model via biochemical approaches. The toxicity of individual mycotoxins on cell viability and cell membrane damage was determined using the MTT and lactate dehydrogenase (LDH) assays, respectively. Individual cytotoxicity of mycotoxins in increasing order were FB1<ZEA<AFB1<DON. The MTT results of central composite design (CCD) showed synergetic effects after co-exposure of AFB1+ZEA or AFB1+DON; however, AFB1 and ZEA showed antagonistic effects in the ternary mixtures. AFB1 and DON significantly induced ROS production and apoptosis in a concentration-dependent manner, but ZEA (10-40μM) had no effect on cell apoptosis and only slightly induced ROS production. ZEA ameliorated the ROS production caused by 1μM AFB1; however, ZEA and DON displayed synergistic effects in combination with AFB1 at 5 and 10μM. The existence of 10μM ZEA attenuated AFB1-induced apoptosis. In conclusion, AFB1+ZEA or DON showed synergetic effects on cytotoxicity. Low levels of AFB1 were antagonistic to ZEA, but high doses of AFB1 displayed synergistic effects with ZEA or DON on oxidative damage. ZEA also ameliorated AFB1-induced apoptosis. Generally, the combined effects of mycotoxins acted in a concentration-dependent manner.

A novel strain of Cellulosimicrobium funkei can biologically detoxify aflatoxin B1 in ducklings.

Two experiments were conducted to screen microorganisms with aflatoxin B1 (AFB1) removal potential from soils and to evaluate their ability in reducing the toxic effects of AFB1 in ducklings. In experiment 1, we screened 11 isolates that showed the AFB1 biodegradation ability, and the one exhibited the highest AFB1 removal ability (97%) was characterized and identified as Cellulosimicrobium funkei (C. funkei). In experiment 2, 80 day‐old Cherry Valley ducklings were divided into four groups with four replicates of five birds each and were used in a 2 by 2 factorial trial design, in which the main factors included administration of AFB1 versus solvent and C. funkei versus solvent for 2 weeks. The AFB1 treatment significantly decreased the body weight gain, feed intake and impaired feed conversion ratio. AFB1 also decreased serum albumin and total protein concentration, while it increased activities of alanine aminotransferase and aspartate aminotransferase and liver damage in the ducklings. Supplementation of C. funkei alleviated the adverse effects of AFB1 on growth performance, and provided protective effects on the serum biochemical indicators, and decreased hepatic injury in the ducklings. Conclusively, our results suggest that the novel isolated C. funkei strain could be used to mitigate the negative effects of aflatoxicosis in ducklings.

Gestational Zearalenone Exposure Causes Reproductive and Developmental Toxicity in Pregnant Rats and Female Offspring

Zearalenone (ZEN) is an oestrogenic mycotoxin commonly found in food and feed products and can affect reproduction and development in both humans and animals. This study aimed to determine the toxic effects of ZEN on maternal SD rats and the F1 female offspring. Sixty-four pregnant rats were divided into 4 groups and exposed to feed contaminated with ZEN (0, 5, 10, and 20 mg/kg feed) on gestational days (GDs) 0–21. Compared with the controls, the groups exposed to 10 and 20 mg/kg ZEN showed significantly decreased feed intake and body weight of pregnant rats and/or female offspring. Meanwhile, 20 mg/kg ZEN significantly decreased the birth weight and viability of F1 newborn rats. Moreover, 10 and 20 mg/kg ZEN diets increased follicle-stimulating hormone concentrations but decreased oestradiol in both maternal and F1 adult rats. In the F1 generation, ZEN caused no pathological changes in ovaries and uterus in weaned rats, but significant follicular atresia and a thinning uterine layer were found in F1 female adult rats in the 20 mg/kg ZEN group. These impairments concurred with the inhibited mRNA and protein levels of oestrogen receptor-alpha (Esr1) and 3β-hydroxysteroid dehydrogenase (HSD) in the adult uterus and/or ovaries. Furthermore, 10 and/or 20 mg/kg ZEN exposure significantly reduced Esr1, gonadotropin-releasing hormone receptor (GnRHr), and ATP binding cassette transporters b1 and c1 (ABCb1 and ABCc1) in the placenta and foetal and weaned F1 brains, and also produced a dose-dependent increase in 3β-HSD in the placenta. Additionally, 20 mg/kg ZEN significantly upregulated ABCc5 expression in the placenta and ovaries of weaned rats. These results suggested that prenatal ZEN exposure in rats affected maternal and foetal development and may lead to long-term reproductive impairment in F1 adult females.

Effects of Nutrients in Substrates of Different Grains on Aflatoxin B1 Production by Aspergillus flavus

The current study was to better understand the potential factors affecting aflatoxin B1 (AFB1) accumulation varies between different grains. The nutrient composition and contents of defatted substrates were determined; additionally, according to the nutrient content of the substrates, the effects of starch, soluble sugars, amino acids, and trace elements on AFB1 production and mycelial growth in Czapek-Dox medium were examined. These results verified that removal of lipids from ground substrates significantly reduced the substrates potential for AFB1 production by Aspergillus flavus. Maltose, glucose, sucrose, arginine, glutamic acid, aspartic acid, and zinc significantly induced AFB1 production up to 1.7- to 26.6-fold. And stachyose more significantly promoted A. flavus growth than the other nutrients. Thus, this study demonstrated that, combined with the nutrients content of grains, in addition to lipids, sucrose, stachyose, glutamic acid, and zinc might play key roles in various grains that are differentially infected by A. flavus. Particularly, two new nutrients (arginine and stachyose) of the grains we found significantly stimulate AFB1 production and A. flavus growth, respectively. The results provide new concepts for antifungal methods to protect food and animal feed from AFB1 contamination.

Aflatoxin B1, zearalenone and deoxynivalenol in feed ingredients and complete feed from central China

ABSTRACT Between 2012 and 2014, 2528 feed ingredient and complete feed samples were collected from central China. Numbers of 2083, 255 and 190 samples were analysed for aflatoxin B1 (AFB1), zearalenone (ZEN) and deoxynivalenol (DON), respectively, by high-performance liquid chromatography in combination with UV or fluorescence detection. The incidence rates of AFB1, ZEN and DON contamination of feed ingredients and complete feeds were 33.9%, 90.2% and 77.4%, respectively. The percentage of positive samples for AFB1 ranged from 13.1% to 97.1%. Cottonseed meal presented the most serious contamination by AFB1. ZEN and DON contamination levels of feeds ranged from 50% to 100%, indicating serious contamination over the studied 3-year period. This study demonstrates that AFB1, ZEN and DON contamination of feeds in central China is serious and differs over the years. Feeds are mostly contaminated with ZEN, followed by DON and AFB1.

Individual and Combined Occurrence of Mycotoxins in Feed Ingredients and Complete Feeds in China

The objective of this study was to investigate the individual and combined contamination of aflatoxin B1 (AFB1), zearalenone (ZEN) and deoxynivalenol (DON) in feedstuffs from different Provinces of China between 2016 and 2017. A total of 1569 samples, including 742 feed ingredients and 827 complete pig feed samples, were collected from various regions of China for mycotoxins analysis. The results showed that individual occurrence rates of AFB1, ZEN, and DON were more than 83.3%, 88%, and 74.5%, respectively, in all the tested samples. DON was the most prevalent contaminant, followed by ZEN and AFB1, with the average concentrations ranging from 450.0–4381.5 μg/kg, 2.3–729.2 μg/kg, and 1.3–10.0 μg/kg, respectively. Notable, 38.2%, 10.8%, and 0.6% of complete pig feeds were contaminated with DON, ZEN, and AFB1 over China’s regulatory limits, respectively. Moreover, over 75.0% analyzed samples were co-contaminated with two or three mycotoxins. In conclusion, the current study revealed that the feedstuffs in China were severely contaminated with DON, followed by ZEN and AFB1 during the past two years. These findings highlight the importance of monitoring mycotoxins in livestock feed and implementing feed management and bioremediation strategies to reduce mycotoxin exposure.

Response of the hepatic transcriptome to aflatoxin B1 in ducklings

This study was conducted to determine the effects of aflatoxin B1 (AFB1) on the hepatic transcriptome in ducklings through RNA-sequencing (RNA-Seq). Twenty four, 1-day-old ducklings were divided into 4 treatment groups. Each group received an oral dose of AFB1 at 0, 10, 20, 40 μg/kg BW per day for 2 weeks. Administration of 20 and 40 μg/kg BW of AFB1 significantly decreased body weight, feed intake, serum total protein and albumin, while increasing serum aspartate aminotransferase and alanine aminotransferase activities, and hepatic histopathological lesions. Furthermore, RNA was extracted from the liver of ducklings administrated 0 and 40 μg/kg BW of AFB1. Two RNA-Seq libraries were created from pooled samples and produced over 149 M reads, totaling 14.9 Gb of sequence. Approximately 96,953 predicted transcripts were assembled, 749 of which had significant differential expressions (≥ 2-fold) between the control and AFB1 treatment. GO and KEGG pathway analysis results showed that many genes involved in phase I metabolism, phase II detoxification, oxidation-reduction process, carcinogenesis, apoptosis and cell cycle, and fatty acid metabolism were affected by AFB1 exposure. Conclusion, this study determined the hepatic transcriptome responded to AFB1 exposure, and provide candidate genes can be targeted to prevent and/or reduce aflatoxicosis in ducklings.

Ameliorative Effects of Grape Seed Proanthocyanidin Extract on Growth Performance, Immune Function, Antioxidant Capacity, Biochemical Constituents, Liver Histopathology and Aflatoxin Residues in Broilers Exposed to Aflatoxin B1

Aflatoxicosis is a grave threat to the poultry industry. Dietary supplementation with antioxidants showed a great potential in enhancing the immune system; hence, protecting animals against aflatoxin B1-induced toxicity. Grape seed proanthocyanidin extract (GSPE) one of the most well-known and powerful antioxidants. Therefore, the purpose of this research was to investigate the effectiveness of GSPE in the detoxification of AFB1 in broilers. A total of 300 one-day-old Cobb chicks were randomly allocated into five treatments of six replicates (10 birds per replicate), fed ad libitum for four weeks with the following dietary treatments: 1. Basal diet (control); 2. Basal diet + 1 mg/kg AFB1 contaminated corn (AFB1); 3. Basal diet + GSPE 250 mg/kg; (GSPE 250 mg/kg) 4. Basal diet + AFB1 (1 mg/kg) + GSPE 250 mg/kg; (AFB1 + GSPE 250 mg/kg) 5. Basal diet + AFB1 (1mg/kg) + GSPE 500 mg/kg, (AFB1 + GSPE 500 mg/kg). When compared with the control group, feeding broilers with AFB1 alone significantly reduced growth performance, serum immunoglobulin contents, negatively altered serum biochemical contents, and enzyme activities, and induced histopathological lesion in the liver. In addition, AFB1 significantly increased malondialdehyde content and decreased total superoxide dismutase, catalase, glutathione peroxide, glutathione-S transferase, glutathione reductase activities, and glutathione concentration within the liver and serum. The supplementation of GSPE (250 and 500 mg/kg) to AFB1 contaminated diet reduced AFB1 residue in the liver and significantly mitigated AFB1 negative effects. From these results, it can be concluded that dietary supplementation of GSPE has protective effects against aflatoxicosis caused by AFB1 in broiler chickens.

Biodetoxification of aflatoxin B1 in cottonseed meal by fermentation of Cellulosimicrobium funkei in duckling diet

&NA; Two experiments were conducted to optimize the fermentation of cottonseed meal by Cellulosimicrobium funkei (C. funkei) for the ability of the bacteria to degrade aflatoxin B1 (AFB1) and then to evaluate the bacterial detoxification in ducklings. In experiment 1, the fermentation of cottonseed meal by C. funkei was improved by changing the inoculation amounts by 10% (108 cfu/mL), using a 1:0.5 material to water ratio at 35°C temperature for a 144 h reaction duration, which resulted in an 83.4% biodegradation of AFB1. In experiment 2, 112 one‐day‐old male Cherry Valley ducklings were randomly allocated to 4 experimental groups with 4 replicates of 7 birds each. For a period of 2 wk the controls received a base duckling diet (BD), a second group received a base diet contaminated with 10% AFB1 cottonseed meal (96.8 &mgr;g AFB1/kg), a third group was fed a base diet added with 5% unfermented and 5% fermented AFB1‐contaminated cottonseed meal (57.0 &mgr;g AFB1/kg), and the fourth group was fed a base diet added with 10% AFB1‐contaminated fermented cottonseed meal (16.0 &mgr;g AFB1/kg). The growth performance, relative organ weights, and serum biochemistry were analyzed. The results showed that the feed conversion ratio in the second group was lower than that of the controls at wk one and 2 (P < 0.05). Also, after 2 wk, group 2 ducklings had increased relative weights of the liver, kidneys, and spleen, increased activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), increased concentration of blood urea nitrogen (BUN) and creatinine (Crt), and decreased relative weight of Fabricius bursa (P < 0.05). In addition, the concentrations of total protein (TP) and albumin (ALB) in serum were also significantly higher at weeks one and 2 (P < 0.05). These alterations were attenuated or prevented when 5 or 10% fermented cottonseed meal substituted equal amounts of unfermented cottonseed meal in the diet. In conclusion, fermentation of AFB1‐contaminated feed materials by C. funkei offers a new strategy to reduce the negative effects of aflatoxicosis in ducklings.

Correction: Rajput, S.A., et al. Ameliorative Effects of Grape Seed Proanthocyanidin Extract on Growth Performance, Immune Function, Antioxidant Capacity, Biochemical Constituents, Liver Histopathology and Aflatoxin Residues in Broilers Exposed to Aflatoxin B1. Toxins 2017, 9, 371

The authors wish to make the following correction to their paper [...].