DNA damage and plasma homocysteine levels are associated with serum metabolites and mineral constituents’ profiles in children with persistent diarrhea

This study describes the association between levels of DNA damage and homocysteine (Hcy) in persistent diarrheic (PD) patients and correlates them with serum biochemical metabolites and mineral components. PD patients (n = 36) age 4 - 6 years from Faisalabad hospitals were examined for anthropometric factors, plasma biochemical and mineral constituents. Compared to 36 normal controls, children with PD had significantly higher concentrations of LDL (p = 0.0001), ALT (p = 0.01),homocysteine (p = 0.001), TOS (p = 0.0001), TBARS (p = 0.001), K (p = 0.0001) and Mg (p = 0.0001) while serum triglyceride, total proteins, albumin, globulin, T3, T4, TAS, Na, Ca, Zn and Cu were significantlylower than those of healthy individuals. Both DNA damage and Hcy were positively linked with LDLcholesterol, TBARS and K (all p values < 0.05). Both Hcy profile and percentage DNA damage in PD patients may impart role in the endothelium damage even in the normal range. PD patients have severe deficiency of macro- and micro-nutrients which may have resulted in enhancement of oxidative stress, DNA damage and Hcy levels in patients’ plasma. Appropriate supplementation of macro- and micronutrients may decrease the DNA damage, Hcy levels and enhance the levels of health markers and decrease the mortality rate of PD patients

Biolistic transformation of Saccharomyces cerevisiae with β-glucosidase gene from Cellulomonas biazotea

A β-glucosidase genomic DNA from Cellulomonas biazotea NIAB 442 was isolated and coated onto tungsten microprojectiles for direct transformation of the gene into Saccharomyces cerevisiae. Transformation of β-glucosidase into S. cerevisae conferred the ability to hydrolyse esculin and cellobiose, indicated that the gene is expressed in the bombarded yeast. Key Words: Biolistic transformation, β-glucosidase, Cellulomonas biazotea, Saccharomyces cerevisiae. African Journal of Biotechnology Vol.3(1) 2004: 112-11

Enhanced production of subtilisin of Pyrococcus furiosus expressed in Escherichia coli using autoinducing medium

A subtilisin gene identified in the reported genome sequence of Pyrococcus furiosus was amplified and inserted in pET-22b(+) vector to produce the recombinant plasmid pET-SB. Escherichia coli BL-21 (DE3)CodonPlus was transformed with this plasmid and the enzyme was expressed up to 30% of the total cell protein on induction with IPTG. The expressed protein appeared at a position corresponding to ~20 kDaon SDS-PAGE as compared to theoretical molecular mass of 17.6 kDa. This aberrant electrophoresis mobility could be due to specific amino acid composition of the protein. Auto-induction with lactose also produced a similar level of expression but the total amount of the enzyme produced was 2.4 foldgreater than that when produced with IPTG induction. This was due to a higher cell density obtainable in the auto-inducing medium. The enzyme expressed in the insoluble state could be partially refolded after denaturation with urea at high pH. This study reports for the first time high-level expression ofsubtilisin of P. furiosus in E. coli using an auto-inducing medium

Influence of nitrogen sources on production of &#946b-galactosidase by Aspergillus niger

The study was undertaken to enhance the production of b-galactosidase using five organic nitrogen sources with wheat bran as a substrate under solid state fermentation. The microbial source Aspergillus niger and its DG-resistant mutant that were grown in medium with initial pH of 5.5 in 250 ml flasks at 30°C for 144 h and sample was harvested after every 24 h and analysed for substrate consumption, cell mass formation and enzyme production. All the nitrogen sources, ammonium sulphate, corn steep liquor, diammonium phosphate, fish meal and urea showed significant results. However, higher values of enzyme activity of 168.0 and 371.15 IU/l/h, parent and mutant, respectively, was obtained from sample in which corn steep liquor was used as a nitrogen source as compared tocontrol (73.1 and 176.3 IU/l/h in parent and mutant, respectively). The effect of nitrogen sources was also found significant in both the organisms but higher in mutant organism (2.2 fold). It is concluded that enzyme production enhanced 2.7 fold by use of suitable production medium under optimum cultural conditions and that the mutant derivative of A. niger can be exploited for hyper production of this enzyme

Infection of hepatitis C virus genotypes in hepatocellular carcinoma patients from rural areas of Faisalabad region, Pakistan

The aim of this retrospective study was to investigate the infection of hepatitis C virus (HCV) genotypes in hepatocellular carcinoma (HCC) patients from rural areas of Faisalabad region. Among 179 HCC subjects, men and women were 51 and 49%, respectively. All samples positive for HCV RNA by qualitative PCR were genotyped, applying genotype-specific PCR. They were confirmed using HCV 5’ non-coding region sequence analytical data. Major risk factor for HCC development and progression was identified to be chronic HCV. It was found among 56.5 ± 2.1 years of age. All these HCC cases were HCV-related and no case was linked with other types of viruses. Using genotype specific primers, HCV genotype 3a (55.3%) was significantly (P<0.0001) higher, followed by 3b (15.8%), 1 (9.24%), 4 (8.05%) and 2 (7.15%). Other genotypes, namely 5a and 6a were only 1.19 and 1.05% among the HCC patients. About 1.05% remained un-typed because of minute viral load. HCV genotype 3a was strongly linked with HCC, followed by 3b. Moreover, HCC was linked with liver cirrhosis (81%). It is suggested that genotyping may be recommended before starting interferon therapy.Key words: Etiology, genotyping, hepatitis C virus, hepatocellular carcinoma, prevalence, α-fetoprotein