Optimization Of Rna Extraction In Mycobacterium Tuberculosis For Studying Intracellular Gene Expression

AOT Alli, JA Mangan, PD Butcher, CL Spreadbury

 

Abstract

Mycobacterium tuberculosis is the leading cause of death due to infectious disease after Human immunodeficiency virus. There has been an upsurge in the incidence of tuberculosis since 1980s. In order to reverse this trend, there is need to understand the biology of the organism. This can be brought about by studying gene expression at transcriptional level. The success of this hinges on RNA of good quality. In this paper, five methods (hot phenol, sonication with guanidinium thiocyanate (GTC) solution, beadbeating method with Trizol, FastPrep machine with Divolab as detergent and GTC solution, and FastPrep machine with Trizol) of extracting RNA from bacteria were compared to find
which of the method would be suitable for mycobacteria. The study found that physical method of lysing bacteria was necessary for extraction of RNA from mycobacteria. FastPrep machine gave the highest yield and also provided the speed necessary for optimum RNA extraction. FastPrep and Trizol as reagent for extraction of RNA was applied to macrophage infected with M. tuberculosis (H37Rv) after removing the macrophage RNA. We were able to demonstrate the expression of dnaK gene in both intracellular and broth grown bacilli. The expression of dnaK gene was found to be downregulated in macrophage compared to broth.

African Journal of Clinical and Experimental Microbiology Vol. 10 (2) 2009: pp. 64-79

Studies On The Distribution Of Clinically Diagnosed Pulmonary Tuberculosis In Ebonyi State, Nigeria

RK Obi, AN Amadi, IM Idika, FC Nwanebu

 

Abstract

A study was conducted between January, 2005 to December, 2006 to ascertain the prevalence of pulmonary tuberculosis among patients who attended chest clinics in some randomly selected hospitals, Clinics and Health Centers in Ebonyi State, Nigeria. Investigations were carried out using the two popular diagnostic criteria for pulmonary tuberculosis namely the specific Ziehl Neelsen (ZN) methods and chest X-ray. A total of 962 patients with clinical signs and symptoms of tuberculosis were studied. Out of this figure, 559 (58.1%) had pulmonary tuberculosis with 1:1 male/female ratio. There was no significant difference between ZN sputum smear positive and chest x-ray in the diagnosis of pulmonary tuberculosis in the studied population at 95% confidence level. The possible reasons for the high prevalence of pulmonary tuberculosis may be attributed to increase in the incidence of HIV/AIDS, high rate of poverty, emergence of drug-resistant strains of Mycobacteria and to a lesser extent, smoking and diabetes

Keywords: Pulmonary tuberculosis, sputum, chest X-ray, Ziehl Neelsen

African Journal of Clinical and Experimental Microbiology Vol. 10 (2) 2009: pp. 80-87

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Sensitivities Of Citrobacter, Proteus And Providencia Isolates To Sulbactam-ampicillin, Trimethoprim-S Ulfamethoxazol And Ticarcillin-Clavulanic Acid Antibiotics

G Uraz, KOK Turhan, HD Binnet

 

Abstract

Gram negative bacterias which belong to Enterobacteriaceae family which is critically important as a matter of human health, are comperatively prevalent in nature and foods. Infections formed by bacterias resistant to antibiotics significantly cause mortality and economical losses. Sensitivities of gram-negative bacterias isolated from miscellaneous samples to sulbactamampicillin, trimethoprim-sulfamethoxazol and ticarcillin-clavulanic acid were analysed. In this study, Proteus is primary isolated microorganism with % 52,08. It was determined that Proteus types are proportionally 86,66% sensitive to Trimethoprim-Sulfamethoxazol and 76% resistant to Sulbactam-Ampicillin . Citrobacter is the secondary isolated microorganism (31,25%). It was determined that Citrobacter types were 86,66% sensitive to Trimethoprim-Sulfamethoxazol and % 73,3 resistant to Sulbactam-Ampicillin. Providencia types which are thirdly isolated microorganism (16%), For Providencia types, Trimethoprim-Sulfamethoxazol were determined as the most effective antibiotic again with 86,66% sensitivity. Resistance to Sulbactam- Ampicillin was however, found to be 76%. In conclusion, it is very important to conduct sensitivity tests in choosing antibiotics for chemotherapy of infections. Trimethoprim-Sulfamethoxazol is recommended in the empiric treatment of urinary tract infections in our environment

Keywords: Enterobacter, sensitive, SXT, TĐM, SAM

African Journal of Clinical and Experimental Microbiology Vol. 10 (2) 2009: pp. 88-91