[Cloning virG gene and generation mutant construct of pGEM delta virG in order to induction recombination in native Shigella dysenteriae]

Shigellosis disease is the major causes of morbidity in children with diarrhea in Iran. The virG [icsA] gene plays a key role in pathogenesis and ability of invasion in shigella. The aim of this study was cloning, sequencing virG gene and developing a mutant construct pGEM delta virG in order to induction recombination in a native shigella for generation a live attenuated vaccine candidate strain. Initially, by use of biochemical tests, the native shigella strain was detected. The virG gene was cloned in pGEM-7zf vector and the nucleotide sequence was determined. According to the data of sequencing, digestion mapping of pGEMvirG vactor was obtained and a part of virG gene by using enzymatic digestion was removed. Finally, pGEM delta virG construct was transformed to E. coli by utilization of chemical transformation method. The native shigella strain by using biochemical tests was confirmed. The result of sequencing virG gene [native strain] was submitted in NCBI Genebank database. The pGEM delta virG construct contains a mutant construct of virG gene which 1751 bp was deleted through enzymatic digestion reaction and transformed in E. coli. Using the technique of allelic exchange based on the incident of recombination in bacteria is one of the most effective methods to develop a disruption in the target genes. This mutant construct can be applied in development of a live attenuated Shigella dysenteriae vaccine candidate.

effect of hyperthyroidism on the levels of liver enzymes in adult male Wistar rats

Thyrotoxicosis is a condition in which tissues are stimulated by increased secretion of thyroid hormone. The most common cause is diffuse toxic goiter and toxic multi-nodular goiter. For more reviews on this disease, the effects of hyperthyroidism on liver enzyme levels were studied. A total of 30 adult male Wistar rats weighing about 190 g were purchased from the Pasteur Institute of Iran. In this study, rats were divided into control group, the group receiving vitamin E, the group receiving levothyroxine, the group receiving levothyroxine treated with vitamin E; blood was taken from all groups over a period of 10 days after injection, and measurement of thyroid hormones and liver tests was made. The findings obtained in this study show that Isolated systolic hypertension [ISH] hormone levels in rats treated with levothyroxine, Treatment with vitamin E may reduce serum levels of ISH, Hormone levels of T4 in the rats treated with levothyroxine were increased compared to normal rates. Treatment with vitamin E reduces serum levels of T4 compared to the first hyper group. T4 hormone levels in rats treated with levothyroxine were reduced compared to normal rates. Treatment with vitamin E may reduce serum levels of T4 compared with the first hyper group. Asparagine Transferase [AST] enzyme levels in rats treated with levothyroxine were increased compared, Treatment with vitamin E may reduce serum levels of AST, Alanine transferase [ALT] enzyme levels in rats treated with levothyroxine were increased, Treatment with vitamin E may increase serum levels of ALT, alkaline phosphatase [ALP] enzyme levels in rats treated with levothyroxine has been increased compared with normal rates. Treatment with vitamin E resulted in serum levels of ALT not to be increased compared with the first group. According to the results of hyperthyroidism and levels of liver enzymes, it can be concluded that hyperthyroidism induced by levothyroxine can increase the levels of hormones T3, T4 and Thyroid Stimulating Hormone [TSH], and then increase the levels of liver enzymes. Treatment of empirical samples with vitamin E is likely to reduce liver damages and prevent the increased levels of liver enzymes compared to empirical samples of hyperthyroidism which have been treated with vitamin E

Detection of sea, seb, sec, seq genes in staphylococcus aureus isolated from nasal carriers in Tehran province, Iran; by multiplex PCR

Staphylococcus[S.] aureus produces different extra-cellular protein toxins and virulence factors. One of the most important extra-cellular proteins is an enterotoxin which causes staphylococcal food poisoning [SFP] due to their enterotoxins. Different methods have been used to detect this toxin, each of which has advantages and disadvantages. DNA amplification methods, however, can show the presence of enterotoxigenic strains of S. aureus before the expression of enterotoxins on the basis of specific gene sequences. In this study, 150 S. aureus strains isolated from nasal carriers were confirmed by biochemical testing. PCR was used to amplify the staphylococcal enterotoxin A, B, C and Q genes, as well as the staphylococcal nuclease gene. Among the 150 healthy human isolates from the nasal carrier, 95 were confirmed as S. aureus. Only 58.9% of the isolates were diagnosed as sea, b, c, q positive. There were 24 [25.3%] isolates associated with the sea gene, 15.8% isolates associated with the seb gene, 9.5% of the isolates were associated with the sec gene, and 8.4% of the isolates associated with the seq gene. Of these isolates, 41% might be possessing additional se genes but they were not see [178 bp] and sed [319 bp] genes. The nuc gene, which encodes thermo nuclease, was used as a target DNA to identify S. aureus. Additionally, one of these enterotoxigenic isolates carried more than one toxin gene

Cloning and Expression of N-terminal Region of IpaD from Shigella dysenteriae in E. coli

Genus Shigella is one of the important members of the family Enterobacteriaceae. There are numerous antigens in Shigella carrying by a 220 kb plasmid. Among them, IpaD is the key virulence factor of S. flexneri. Apart from having effectors function that is essential for host cell invasion and intracellular survival, this protein also controls the secretion and translocation of other effector proteins into eukaryotic host cells. In the present study, we have cloned and expressed the ipaD in E. coli. The ipaD gene was amplified by PCR. Prokaryote expression vector pET-28a [+] - ipaD was constructed, and used to transform E. coli BL21DE3 plySs. The expression of recombinant protein induced by IPTG was examined by SDS-PAGE. Western blot were used to determine immunoreactivity of IpaD-His by a rabbit monoclonal antibodies against his-tag. SDS-PAGE demonstrated that the constructed prokaryotic expression efficiently produced IpaD at the 1 mmol/L of IPTG. IpaD protein was able to react with the rabbit monoclonal antibody against His-tag. IpaD is essential for Shigella spp invasion. N-terminal region is most significant functional fragment of IpaD. Purification of IpaD from the wild type of Shigella is difficult furthermore profound study on a specific domain on the N-terminal of IpaD by using the wild type of purified IpaD is not feasible