Genomic study of the cereolysin A and B genes in Bacillus cereus isolated from raw and pasteurized milk

Bacillus cereus spores are expanded in the nature and they can be separated from different foods. Cytotoxin is one of the most important poisons which is produced by Bacillus cereus, and it is highly resistant to heat and leads to diarrhea, nausea and vomiting syndrome. Hence study about existence of Bacillus cereus in pasteurized milk is very important due to probability of causing illness by Cereolysin gene products. Therefore, Different milk samples were collected from raw milk to pasteurized milk after various stages of producing pasteurized milk. Cultivation of milk samples in Mannitol egg yolk polymyxin B (MYP) media was done and it was followed by a purification of the observed colonies andinvestigation of cereolysin A and B by polymerase chain reaction (PCR) amplification. Thus, the gene was amplified and aligned with the previous sequences that were registered in the gene bank database. As such, many new missense mutations were observed at the amplified gene sequences. However these missense mutations caused a change in the sequence of amino acid at the protein chain but the protein efficiency and structure was not changed due to the substitution of amino acids with the sameproperties.Key words: milk- cereolysin - Bacillus cereus- polymerase chain reaction- gene

Optimization of in vitro culture and transfection condition of bovine primary spermatogonial stem cells

The present study aimed to optimize the in vitro culture and transfection efficiency of bovine primary spermatogonial stem cells (SSCs). To this end, SSCs were obtained from newborn Holstein bull calves by two-step enzymatic digestion. After enrichment and culture, SSCs were characterized by using alkaline phosphatase (AP) staining and expression of vasa and thy1 genes as specific bovine SSC markers. To evaluate the effect of antioxidants on vitality, colony formation, and the expression of pro- and anti-apoptotic genes of bovine SSCs, various concentrations of vitamin C (5, 10, 25 and 50 μg/mL) and Trolox (a water soluble α-tocopherol analogue) (12.5, 25, 50 and 100 μg/mL) were added to the SSC culture medium. The results showed that SSCs treated with 50 μg/mL of vitamin C or 25 μg/mL of Trolox individually could increase cell viability and colony formation significantly in comparison with other concentrations and the control group. Additionally, the expressions of bax (as a pro-apoptotic gene) and bcl2 (as an anti-apoptotic gene) were significantly lower and higher than the control group, respectively. To optimize the transfection condition, the effective dosages of vitamin C or Trolox, with various concentrations of two transfection reagents (X-tremeGENE HP and Turbofect) and DNA, at day 8 of culture, were studied. Results showed that 1 μl X-tremeGENE HP or 0.5 μl Turbofect and 2 μg of DNA are the best concentrations for transfecting SSCs. However, X-tremeGENE HP expressed more potential for transfecting SSCs in comparison with Turbofect. Besides, no difference was observed between the use of defined doses of vitamin C or Trolox.Keywords: Apoptosis, gene transfer, primary cells, viability, vitamin C, Trolo

Growth performance, carcass yield and intestinal microflora populations of broilers fed diets containing thepax and yogurt

The present study aimed at evaluating the effect of the probiotic thepax and yogurt (as probiotic) on the growth response and intestinal microflora results of broiler chickens. Two hundred forty day-old Ross 308 broilers were equally distributed into 12 floor pens and reared for 42 day. The treatments consisted of yogurt (10, 5 and 2.5% during starter, grower and finisher periods in the drinking water, respectively) and thepax (1000, 500, 250 g/ton-1 in the starter, grower and finisher diets, respectively), resulting three experimental diets and a control group. Each dietary treatment was fed ad-libitum to four replicate group of 20 birds at the beginning of rearing period. Birds and feed were weighed on days 21 and 42. The results of experiment indicate that diets containing feed additives improved broiler performance. The body weight gain and feed conversion ratio improved significantly more (p < 0.05) with the thepax treatment compared with the control broilers during the total rearing period. The highest (p < 0.05) carcass and thigh values were recorded for broilers fed the diet supplemented with thepax and yogurt, respectively. The lowest abdominal fat pad value was obtained in broilers fed the diet supplemented with thepax. On d 21, thepax and yogurt significantly reduced (p < 0.05) cecal Escherichia coli and Clostridium perfringens populations compared with the control group. In conclusion, thepax and yogurt improved broilers growth response and conferred intestinal health benefits to chickens by improving their microbial ecology

Carcass Traits and Immune Response of Broiler Chickens Fed Dietary L-Carnitine, Coenzyme Q10 and Ractopamine

ABSTRACT This study was conducted to evaluate the effects of coenzyme Q10, L-carnitine and ractopamine supplementation, alone and in combinations, on carcass traits and immune response of broiler chickens. Five hundred and twelve one-day-old Ross 308 male broiler chickens were randomly allocated into eight treatments with four replicates each. A 2×2×2 factorial arrangement was applied, with two levels of coenzyme Q10 (0 and 40 mg/kg), two levels of L-carnitine (0 and 200 mg/kg) and two levels of ractopamine (0 and 10 mg/kg). The birds were reared until day 42 of age under standard conditions. Blood samples were collected at the end of grower and finisher periods from the wing vein. Four birds per group were sacrificed at day 42 of age. Except for carcass yield, other carcass traits were not significantly affected (p>0.05) by different levels of coenzyme Q10, L-carnitine, or ractopamine. Immune response parameters were significantly (p<0.05) different between the treatments. The lowest antibody titers against Newcastle disease virus and relative spleen weight were observed in control group. The results of this study suggest that addition of coenzyme Q10 and L-carnitine to broiler diets has benefit effect on immune response of broiler chickens

Evaluating bovine sperm transfection using a high-performance polymer reagent and assessing the fertilizing capacity of transfected spermatozoa using an in vitro fertilization technique

Sperm-mediated gene transfer (SMGT) has been considered as an innovative device for transgenesis on a mass scale by taking advantage of live spermatozoa to transfer exogenous DNA. However, the fertilizing ability of transfected sperm cells and the poor reproducibility of this method are still matters of controversy. Hence, the current study was conducted to evaluate transfecting the enhanced green fluorescent protein (EGFP) as the source of exogenous DNA into bovine spermatozoa using a high-performance polymer reagent as well as assessing the fertilizing capacity of transfected sperm cells by in vitro fertilization (IVF). In the first experiment, three different concentrations of rhodamine-labeled DNA and high-performance polymer transfection reagent, X-tremeGENE HP, were used to transfect bovine spermatozoa. In the second experiment, IVF and fluorescence microscopy methods were utilized to assess the fertilizing capacity of sperm cells carrying exogenous DNA when X-tremeGENE HP was used either alone or with dimethyl sulfoxide (DMSO) treatment. Findings revealed that at 1&thinsp;µL X-tremeGENE HP and 1&thinsp;µg of DNA concentration, approximately one-third of total spermatozoa were transfected. However, following IVF and fluorescence microscopy, no EGFP expression was detected in zygotes and morula-stage embryos. Results of this study showed that, although X-tremeGENE HP could transfer EGFP to bovine spermatozoa, transfected sperm cells were unable to transfer foreign DNA to matured bovine oocytes. Under our experimental conditions, we hypothesized that the absence of the EGFP fluorescence signal in embryos could be due to the detrimental effects of transfection treatments on sperm cells' fertility performance as well as incompetency of IVF to produce transgenic embryos using transfected sperm cells.</p