Use of enzyme linked immunospot assay [ELISpot] for monitoring treatment response of pulmonary tuberculosis patients

Tuberculosis [TB] remains one of the major causes of death from a single infectious agent worldwide. The rapid emergence of drug resistant mycobacteria has strengthened the demand for rapid methods for detection of mycobacteria in clinical samples. As prevention of tuberculosis relies on the early detection and cure of the infectious cases, current efforts are focused upon improving the rapidity of identification of Mycobacterium tuberculosis, allowing prompt initiation of appropriate therapy. The rapid enzyme linked immunospot assay [ELISpot] method was developed in the late 1990s based on the numbers of spots made by interferon gamma producing T cells stimulated by culture filtrate protein-10 [CFP-10] or early secretory antigenic target-6 [ESAT-6]. Therefore, a T-cell response to these antigens could in theory serve as a specific marker of M. tuberculosis infection. Is to assess the potential utility of ELISpot assay for monitoring treatment response of pulmonary tuberculosis patients. The study was done on 30 patients diagnosed as pulmonary tuberculosis on clinical, radiological and bacteriological bases. They were collected from Zagazig Chest Hospital and Zagazig University Hospitals from January 2010 to January 2011. A total of 15 healthy volunteers were enrolled in this study as control subjects. The following were performed for all patients before treatment initiation: full history taking, complete clinical examination, chest X-ray, postero-anterior and lateral views, tuberculin skin test [TST] by Mantoux technique, and routine laboratory investigations. Three successive sputum samples for sputum smear Ziehl-Neelsen [Z-N] staining and sputum collection for Mycobacterium culture on Lowenstein-Jensen media [LJ. Media] were done. Collection of 2 ml heparinized blood for enzyme linked immunospot assay [ELISpot] was done. All patients received four antituberculous drugs, isoniazid, rifampicin, pyrazinamide and ethambutol, for the initial 2 months. After 2 months of therapy; another three successive sputum samples for sputum smear Ziehl-Neelsen [Z-N] staining and sputum collection for Mycobacterium culture on Lowenstein-Jensen media were done. Collection of 2 ml heparinized blood for enzyme linked immunospot assay [ELISpot] was done. The results of this study showed that all patients were complaining of cough and expectoration. Tuberculin skin test was positive in 18 patients [60%]. Most patients [46.6%] had moderately advanced disease as regards the radiological extent. It was found that the median INF-y ELISpot response to ESAT-6 was significantly decreased after 2 months of antituberculosis therapy. The number of pre-treatment ESAT-6 ELISpot count in patients with positive tuberculin skin test was significantly higher than those with negative tuberculin skin test [P < 0.01]. As regards bacillary load, a statistical significant difference between patients with AFB+ + + and patients with [AFB +, AFB + +] as regards pre-treatment ELISpot count was recorded. Higher statistical significant difference in patients with AFB +, AFB + + and AFB + + 4- pre and post treatment ESAT-6 ELISpot count was found. It was found that the number of pre-treatment ESAT-6 ELISpot count in patients with cavitary lesion was higher than those without cavitary lesion and the difference was highly significant [P = 0.01]. As regards radiological extent, it was found that the number of pre-treatment ESAT-6 ELISpot count in patients with far advanced disease was higher than patients with minimal or moderately advanced disease. Also, after 2 months of therapy the number of ESAT-6 ELISpot count in patients with far advanced disease showed more decline than patients with minimal or moderately advanced disease. It was found that ELISpot assay sensitivity, specificity, positive predictive value and negative predictive value in relation to L.J. media were 93.3%, 100%, 100% and 88.2%, respectively. ELISpot assay may be used as a useful tool in the diagnosis of pulmonary tuberculosis. The decrease in the M. tuberculosis-specific T cell responses following 2 months of successful antituberculosis therapy may have a clinical value as a supplemental tool for the monitoring treatment response of pulmonary tuberculosis patients

Extended spectrum B-lactamase-producing Escherichia coli in clinical isolates in Benghazi, Libya: phenotypic detection and antimicrobial susceptibility pattern

beta-lactams are the most widely used group of antimicrobials; however, growing resistance to these invaluable drugs mediated by extended spectrum beta-lactamase [ESBL] enzymes is a major concern. The present study was undertaken to determine the prevalence of these enzymes and their effect on antimicrobial susceptibility pattern by different phenotypic detection tests in clinical isolates of Escherichia coli in Benghazi, Libya. Antimicrobial susceptibility testing was carried out by Kirby-Bauer method. Ceftazidime and cefotaxime were used for screening potential ESBL producers. Confirmation was done by a combination of double disk synergy test [DDST] and phenotypic confirmatory disk diffusion tests [PCDDTs]. A total of 120 E. coli strains [40 urine, 20 sputum, 20 blood, and 40 wound swabs] from inpatients at different hospitals of Benghazi, Libya, were included in the study, of which, 24 [20%] isolates were ESBL producers. The resistance pattern to the tested antibiotics was as follows: ampicillin [80%], co-trimoxazole [60%], ciprofloxacine [40%], cefotaxime [30%], ceftazidime [30%], Ceftriaxone [30%], gentamicin [30%], cefpirome [35%], ofloxacin [30%], imipenem [25%], and nitrofurantoin [40%]. All the isolates tested showed resistance to two or more drugs and were considered to be multidrug resistant. A higher rate of ESBL production and multidrug resistance was seen among isolates from pus swabs as compared to other sources. ESBL producers mediated high resistance to both beta-lactams and non-beta-lactams. Prolonged hospital stay and prior use of third-generation cephalosporins were identified as important risk factors for ESBL acquisition. There is insufficient data regarding ESBL prevalence among E. coli strains from Benghazi, Libya. ESBLs not only pose a great threat to future of beta-lactams, but they also endanger the utility of many non-beta-lactams. To ensure rationale in antibiotic treatment, ESBL detection and reporting assumes a priority in near future in Benghazi, Libya

Prostate-specific antigen versus digital rectal examination in the diagnosis of prostate cancer

Prostate cancer [CaP] is the most commonly diagnosed non cutaneous cancer and the third most common cause of death from cancer in males. PSA is considered the most useful tumor marker currently available for diagnosis and management of the CaP. The digital rectal examination is still the basis in the suspicion of CaP in males with normal or minimally high PSA levels. The aim of the study is to evaluate the effectiveness of PSA versus digital rectal examination [DRE] in identifying cases of prostate cancer among men presenting with symptoms of bladder outflow obstruction. A cross sectional study was conducted at Azadi Teaching Hospital in Dohuk on 400 patients with bladder outflow obstruction above 50 years were selected between January 2005 and October 2007. DRE and serum PSA done for all patients and prostate biopsy for abnormal results. The mean age 69.63 +/- 8.3 years. All cases of documented prostate cancer with positive DRE had PSA >/= 4 ng/ml. In conclusion, DRE is still the basic step in the suspecting prostate cancer. Combination of DRE and PSA has the highest detection rate for CaP than each alone

Epstein-BARR virus in Iraqi patients with nasopharangeal carcinoma

Epstein-Barr virus [EBV] was ubiquitous Herpes virus that had a role in the development of undifferentiated carcinoma of the nasopharynx, Burkett's lymphoma, acute infectious mononucleosis and other lymphoprolifrative disorders. Thirty Iraqi patients with nasopharangeal carcinoma were referred to Oncology Unit in Al-Kadhemia Teaching Hospital from 1992-1994. Sera of those patients were tested for the presence of antibodies against Epstein-Barr virus nuclear and early antigens using indirect immunoflourescence test. Cellular immunity for those patients was tested for the CD4+, CD8+, CD4/CD8 ratio, T-cells% and B-cells%. Their results were compared with twenty-two normal apparently normal individuals. Antibodies to Epstein-Barr virus nuclear and early antigens were detected in nasopharangeal carcinoma Iraqi patients and not in the control group. There was significant difference between two groups in CD8+cells, T-cells% and B-cells% and there was no significant differences between two groups in CD4+cells, CD4/CD8 ratio. EBV infection was stopped by T-cells immune response that was capable of eliminating virus infected cells and virus neutralizing antibodies against nuclear and early antigens which prevent the spread of infection. Lymphocytes were predominantly CD8+cytotoxic T lymphocytes, which recognize and destroy EBV infected cells. Other antibodies to viral capsid antigens [IgG, IgA and IgM]. Other methods must be used other than indirect immunoflourescense test like western blot method and enzyme linked immune sorbent assay [ELISA]

Prevalence of autoantibodies to various tissue antigens before and during S2-complex immunotherapy in head and neck cancer patients

A total of 63 terminal untreatable stage IV head and neck cancer patients were investigated for clinical responses and presence of autoantibodies to various tissue antigens before and during S2-complex immunotherapy. S2-complex is a new low molecular weight biological response modifier [BRM] with a potant immunostimulating and anti tumor activities. Autoantibodies detected at pretreatment period were those directed towards the following antigens nuclear, thyroid microsomal, epithelial cells, gastric parietal cells, smooth muscles, peripheral leukocytes, T-lymphocytes, B-lymphocytes, monocytes, thymus reticular epithelial cells and Hassall's corpuscles. Beside, autoantibodies with specificities to glomerular basement membrane and vascular endothelial cells were present at low incidence. Short term use of S2-complex induced a transient increase of the following autoantibodies: nuclear, thyroid microsomal, epithelial, parietal cells, smooth muscle, thymus reticuloepithelial cells, Hassal's corpuscles, thymocytes, peripheral blood leukocytes, 1, B-lymphocytes, monocytes, as well as kidney glomerular basement membrane and vasculr endothelial cells. In the later follow up period i.e. 2-6 months most of these autoantibodies responses returned to normal healthy control levels. Moreover, two exceptions were demonstrated which were the incidences of the antiglomerular basement membrane and vascular endothelial antibodies which remianed higher than the pretreatment frequencies. In addition, autoantibodies specific to mitochondria, thyroglobulin and red blood cells were only occasionally seen in our head and neck cancer patients both before and during S2-complex therapy

P53/WAF1 dependent pathway in squamous cell carcinoma of bladder associated with Bilharziasis

Our understanding of cellular growth control and cell cycle control has been studied extensively at the last decade. Several genes have been identified whose expression are affected by p53 gene. The waf1 is a target gene whose expression has been found to be regulated by p53. We studied p53/waf1 expression inschistosomal egyptian bladder cancer patients in an attempt for the possible use of them as genetic marker for assessment of the progression of such type of disease in egypt. For this purpose, 50 bladder tumors and their corresponding safty margins as controls were collected from nci, cairo niversity and screened for detection of p53 alterations using western blotting confirmed by pcr-sscp analysis, as well as the expression of waf1 gene using western blotting technique. Our results revealed that 20 cases [40%] showed mut-p53, 19 of them harbor abnormal pattern of sscp analysis. A significant correlation was reported between p53 and histopathological type of bladder cancer [scc and tcc, p=0.02]. We reported 21 cases [42%] showed waf1 downexpression. This study gives a dramatic example for the interaply between the oncosuppressor genes in cancer where, a very strong negative association was reported between the presence of p53 alteration [at both dna and protein levels] and waf1 expression [p=0.001]. In conclusion, this study provides an evidence that p53 may be involved in scc genetic. However, no role for p53 alterations in the progression or metastatic potential of bladder cancer. Also, p53/waf1 deficiency may not involve in bladder cancer development and/or progression