- A-Z Publications
- Molecular Diagnosis and Vaccines
- OA African Journal Archive
- Volume 1, Issue 1, 2003
Molecular Diagnosis and Vaccines - Volume 1, Issue 1, 2003
Volume 1, Issue 1, 2003
Sequencing emm gene for accurate typing of Egyptian-hemolytic streptococci isolates from non-sterile sitesSource: Molecular Diagnosis and Vaccines 1, pp 1 –9 (2003)More Less
Group A streptococcal (GAS)-mediated diseases continue to be a major problem world wide. Typing of clinical isolates of GAS relies primarily on serologic typing of surface M Protein using polyclonal antibodies. In this study, problems associated with M serotyping such as limited availability of typing sera, newly encountered M types and difficulty in interpretation were avoided by using a system based on sequence analysis of the emm gene that encode M serospecificity. This method relies upon the use of two highly conserved primers that amplify a large portion of the emm gene and the fact that the hyper variable sequence encoding M serospecificity lies adjacent to one of the amplifying primer sequence allowing direct sequencing. One hundred and forty ?-hemolytic streptococci isolates from non sterile sites (94 impetigo, 46 pharengeal) of Egyptian patients were subjected to emm typing. Forty five emm types were identified, 12 of them never been encountered in USA or Egypt and 6 out of these 12 emm type strains were newly discovered worldwide. In addition, several new emm-T-agglutination pattern-associations were found. These findings indicate that Egypt has a unique distribution of group A streptococci strains.
Source: Molecular Diagnosis and Vaccines 1, pp 11 –19 (2003)More Less
This study was done to investigate the role of some risk factors for development of peripheral neuropathy in children and young adolescents with type I diabetes mellitus (type I DM), Diabetic patients were divided into three groups: Group I included 10 diabetic patients with clinically and electrophysiologically evident peripheral neuropathy, Group II included 20 diabetic patients with subclinical peripheral neuropathy evident only by measuring the motor nerve conduction velocity (MNCV) and Group III included 30 diabetic patients without any evidence of peripheral neuropathy. Ten normal healthy individuals with matched age and sex served as a control group (Group IV). All the studied groups were subjected to full clinical and neurological examination, measuring (MNCV) of the common peroneal and median nerves, estimation of glycosylated hemoglobin level (HbA1c), assay of erythrocyte superoxide dismutase (SOD) level as well as molecular genetic study of the manganese superoxide dismutase (Mn-SOD) gene polymorphism. Our results showed that there was a significant increase in HbA1c level and a significant decrease in both erythrocyte SOD level as well as a significant decrease in MNCV of common peroneal and median nerves in group I and group II as compared with group III and group IV. There were significant negative correlations between HbA1c when compared with SOD and MNCV as well as significant positive correlation between SOD levels and MNCV in all the diabetic groups. The frequencies of Ala allele and the Ala/Ala genotype of the Mn-SOD gene were significantly lower in neuropathic diabetic patients.In contrast, the Val allele and Val/Val genotype were significantly more frequent in neuropathic diabetic patients than in diabetic subjects without peripheral neuropathy. This suggests that the Ala(-9)Val dimorphism in the Mn-SOD gene is associated with neuropathy in type 1 diabetes mellitus. In conclusion, Poor glycaemic control, low levels of the key antioxidant enzyme SOD, and the Ala(-9)Val genotype of the Mn-SOD gene were significant risk factors for the development of diabetic neuropathy in type 1 diabetic patients which may necessitate appropriate support for enhancing antioxidant supply to act against the rapid onset and progression of peripheral neuropathy where as the Ala allele and Ala/ Ala genotype are associated with low risk of neuropathy in patients with type 1 DM.
Soluble P-selectin level as a predictor of prothrombotic status in patients with atrial fibrillationSource: Molecular Diagnosis and Vaccines 1, pp 21 –28 (2003)More Less
The increased risk of thromboembolism in atrial fibrillation (AF) may be related to a prothrombotic or hypercoagulable state, with abnormalities of hemostasis and platelet activation. P-selectin is an adhesion molecule that is expressed by platelets on activation and soluble P-selectin (sP-selectin) level is now considered a reliable marker of in vivo platelet activation. We investigated the value of sP-selectin level as a marker of prothrombotic state in patients with AF. Thirty patients with documented permanent AF (16 males, 14 females, mean age 5611.8 years) were subjected to transthoracic (TTE) and transesophageal (TEE) echocardiographic studies. sP-selectin level was measured using ELISA in these patients and in 12 age matched healthy controls. Results revealed a significant elevation of sP-selectin levels in patients with AF as compared to control group (16139 ng/ml vs. 16.7 8.9 ng/ml, respectively), (P<0.001), in spite of proper anticoagulation. In the patient group, significant elevation in sP-Selectin levels was noted in patients with diabetes mellitus (17639 ng/ml), as compared to non-diabetic patients (150.936.6 ng/ml) (P<0.05). The presence of spontaneous echo contrast (SEC) or left atrial thrombus in TEE was associated with significant elevation of sP-selectin levels, as compared to those of patients having none of these findings (179.8 36.8 ng/ml vs. 130.523.1 ng/ml, respectively), (P<0.01). Platelet activation occurs as part of the prothrombotic status in patients with AF in spite of proper anticoagulation. Determination of sP-selectin level may provide a useful noninvasive marker to identify AF patients at high thromboembolic risk.
Circulating CD4+ and CD8+ T-Cell profiles in pulmonary tuberculosis: association with disease activity and response to treatmentSource: Molecular Diagnosis and Vaccines 1, pp 29 –38 (2003)More Less
The wide spectrum of clinical outcomes following infection with Mycobacterium tuberculosis (Mtb) is largely determined by the host immune response. This tudy assesses circulating CD4+ and CD8+ T-cell profiles in patients with pulmonary tuberculosis in relation to disease activity before and 2 months after the start of antituberculous chemotherapy. Two-colour Flow Cytometry was used for counting circulating T-cell subpopulations. Forty patients with pulmonary tuberculosis and 10 age and sex matched normal controls were involved in this study. Patients were divided according to the severity of disease into 28 patients with minimal or moderate disease and 12 patients with severe extensive disease. Sputum and blood specimens were obtained at admission and 2 months after the start of treatment. Although, no significant differences in CD4+ T- cell counts (mean %) between normal controls and both patients with mild/moderate disease and with severe disease at admission and 2 months after the start of treatment (P>0.05) yet, significant differences were observed in CD8+ T-cell counts between normal controls and both patients with mild/moderate and severe disease at admission (40.78 8.60) and (46.16 6.01) respectively and 2 months after the start of treatment only in patients with severe disease (46.00 5.15) (P<0.001). At admission a significant difference (P<0.05) was observed between patients with mild/moderate and severe disease only in CD8+ T-cell counts while after 2 months of the start of treatment, there were significant differences in both CD4+ (48.78 8.87 and 43.16 6.22) and CD8+ (35.35 37.35 and 46.00 5.15) respectively. In conclusion, circulating CD4+ and CD8+ T-cell profiles change with disease activity and drug efficacy. The study suggests the possible use of CD8+ T-cells in prediction of disease outcome and monitoring drug efficacy and may point out to the potential role of CD8+ T-cells in the development of a novel vaccine strategy against tuberculosis.
Methylene tetrahydrofolate reductase gene (C677T) polymorphism, folic acid, vitamin B12 and homocysteine in coronary heart diseaseSource: Molecular Diagnosis and Vaccines 1, pp 39 –47 (2003)More Less
Hyperhomocysteinemia, is a risk factor for occlusive arterial disease. It may be caused by disruptions of homocysteine metabolism. The C677T mutation in methylenetetrahydrofolate reductase (MTHFR) gene may contribute to elevated plasma homocysteine and is probably one of the atherosclerotic risk factors. This work was carried out to investigate the relationship between the MTHFR C677T mutation, plasma homocysteine concentration and the risk of coronary artery disease (CAD) in 48 angiographically confirmed CAD male patients. Control group comprised 20 age-matched healthy subjects. The MTHFR gene mutation was detected by PCR and restriction digestion. Plasma homocysteine, Folic acid and vitamin B12 were assayed by a chemiluminescence method. The incidence of the mutated T allele and genotype CT was similar in patients and controls (0.156 vs 0.125, and 31.25% vs 25% respectively, p = 0.6). No TT homozygotes were detected. Homocysteine was significantly higher and folic acid signicantly lower in patients than controls (p = 0.0004 and <0.0001 respectively). Although CT heterozygotes had significantly higher homocysteine concentration than homozygous CC subjects in both patients and controls (p<0.0001), the odds ratio for CAD in genotype CT was 1.3 (p = 0.6, NS). The odds ratio for CAD in subjects with hyperhomocysteinemia was 7.6 (p = 0.01). After multivariate analysis, hyperhomocysteinemia remained an independent risk factor of CAD (OR 5.7, p = 0.04). in conclusion, hyperhomocysteinemia rather than the C677T mutation in the MTHFR gene, is an independent risk factor of coronary artery disease.
Molecular diagnosis of mycoplasma pneumoniae in children with atypical pneumonia: comparison with the standard culture procedureSource: Molecular Diagnosis and Vaccines 1, pp 49 –57 (2003)More Less
Mycoplasma pneumoniae causes a variety of respiratory tract infections in young children and adults. The standard laboratory methods for its diagnosis are culture and serology, but this agent being fastidious and growing slowly limits the usefulness of culture for routine purposes. Polymerase chain reaction (PCR) technique has recently been used as a rapid method for diagnosis of M. pneumoniae infection. In this study we compared culture on selective media as a standard method and direct detection of M. pneumoniae DNA in clinical specimens by PCR. Throat specimens from 40 children suffering from atypical pneumonia and 20 apparently healthy children were studied. A significant difference was found between atypical pneumonia cases and controls as regard total leucocytic count and erythrocyte sedmintation rate. Eight cases (17.5%) were culture positive, while 12 (27.5%) were PCR positive. All culture positive cases were PCR positive, PCR sensitivity was 100%, specificity 92% and accuracy 93.3%. In conclusion, PCR is a sensitive and rapid method for diagnosis of M. pnemoniae infection, while culture on selective media although being specific but time consuming. The high sensitivity of the test allows early initiation of proper antibiotic therapy to decrease morbidity of the disease.
Source: Molecular Diagnosis and Vaccines 1, pp 59 –70 (2003)More Less
Pseudomonas aeruginosa is an important nosocomial pathogen, especially in individuals who are immunocompromised. This study investigates pseudomonas infections in burn wounds of patients at the Menoufiya University Hospital. Pseudomonas isolates were identified to the species level using automatic Sensititre, subjected to antibiotic susceptibility testing. Pseudomonas aeruginosa was the predominant (56.5%) species of Pseudomonas while mixed excellent group, Pseudomonas putida and Pseudomonas fluorscens constituted 19.6%, 15.2% and 8.7% of the isolates respectively. All Pseudomonas isolates were resistant to sulphamethoxazole trimethoprim (SXT), amoxycillin (AML), ampicillin + sulbactam (SAM), cefepime (CP), tetracycline (TE). Most of the strains were resistant to erythromycin (E) (91.3%), amoxycillin + clavulanic acid (AMC) (74.9%), ceftriaxone (CRO) (63%), azetreonam (ATM) (56.5%).Most of the strains were sensitive to amikacin (AK), tobramycin (TOB), gentamicin (CN), cefoperazone (CFP), and piperacillin (PRL) (91.3%, 89.2%, 69.6%, 58.7%, 56.6%) in this descending order of activity. Twenty isolates (43.4%) were ? lactamase producer, out of these isolates, 16 strains (80%) contained plasmids, while 4(20%) were plasmidless. Extended spectrum ? lactamase (ESBLS) was demonstrated in 4(8.6%) of isolates. Plasmid profile analysis showed that 18 isolates (39.1%) harboured plasmids with molecular weight ranging from 1.4 140 MDa. The number of plasmids, ranged from one to five, 8 strains (44.4%) contained 3 plasmids while 7 (38.9%) had one plasmid. 2(11.1%)had 2 plasmids, and one (5.5%) contained 5 plasmids. Resistances to AK, TOB and ATM were associated with the existence of large molecular weight plasmids. Moreover, resistance to increasing number of antibiotics was associated with high molecular weight plasmids. In conclusion, the burn wound represents a susceptible site for opportunistic colonization and multidrug resistant strains, Pseudomonas aeruginosa infection is particularly problematic, plasmid may be responsible for resistance to antibiotics. Resistance to ESBLS is an important problem in Egypt. Implementation of infection control measures may help in reducing further transmission of Pseudomonas clones within the burn units.
Evaluation of different methods of identification of methicillin-resistant Staphylococcus aureus (MRSA); antibiogram and plasmid profile of hospital and community-acquired strainsSource: Molecular Diagnosis and Vaccines 1, pp 71 –85 (2003)More Less
Nosocomial infections by methicillinresistant Staphylococcus aureus (MRSA) represent an increasing problem in hospitals. The aims of this study were to assess the prevalence of MRSA in Shebin El-Kom Teaching Hospital, to evaluate oxacillin screen agar, oxacillin disc diffusion and E-test in the identification of MRSA, to analyze the plasmid profile and know the role of hospital staff in transmition of infection within the hospital. The results of the present study showed that Staph. aureus constituted 40.2% of all nosocomial infections and that 76 out of the isolated 177 Staph. aureus strains were resistant to oxacillin (66 MRSA strains were isolated from 132 inpatients, 7 from 25 outpatients, and 3 from 38 medical staff). All isolated MRSA strains were multi-drug resistant with a high rate of resistance to penicillin, ampicillin, oxacillin, amoxicillin-clavulanate, cephalexin, ceftriaxone, erythromycin, tetracycline and gentamycin. On the other hand, they were mostly sensitive to rifampicin, clindamycin, vancomycin and teicoplanin. Oxacillin screen agar base (ORSAB) was more sensitive as compared to both E- test and oxacillin disk diffusion in detecting MRSA (76 MRSA isolates were identified by ORSAB while 73 and 72 MRSA strains were detected by the Etest and disc diffusion respectively). Plasmid profile analysis showed that 55 (72.4%) MRSA strains harboured plasmids, while 21(27.6%) were plasmidless. The molecular weight of plasmids ranged from 1.2 to 23 MDa and the number of plasmids per isolate ranged from 1 to 5. Nine out of 27 MRSA strains isolated from clinical sites contained a 1.4 MDa plasmid which was occasionally present in strains isolated from medical staff who had an antibiotic resistance pattern which was similar to patients strains. In conclusion, multi-drug resistant MRSA strains are important cause of nosocomial infection. Most of them contained plasmids which may be responsible for antibiotic resistance. Oxacillin screen agar may be a reliable and rapid method for identification of MRSA. Surveillance of medical staff for MRSA carriage may be helpful in detecting carriers and subsequent better control of MRSA spread.
Relationship between fetal and neonatal cardiac functions and degree of glycemic control in pregnant diabetic womenAuthor Maged Ramses FahimSource: Molecular Diagnosis and Vaccines 1, pp 87 –94 (2003)More Less
Aim of work was to assess the effect of various degrees of diabetic control during pregnancy on the cardiac function in fetuses and infants at 24-72 hours after birth. Echocardiographic examination of fetuses was done at 6 weeks intervals starting at 20 weeks of gestation in 12 fetuses of good control diabetic women, 5 fetuses of fair control diabetic women and 3 fetuses of poor control diabetic women. Echocardiographic examination was reported at 24-72 hours after birth. Glycosylated hemoglobin of the mothers was estimated early in pregnancy at 8 weeks gestation and then at 8 weeks intervals. Maternal diabetic control was classified into good, fair and poor control according to the maternal Glycosylated hemoglobin levels. Twenty age matched healthy nondiabetic pregnant women and also 20 age and sex matched normal healthy newborns of comparable gestation and birth weight were used. In good control diabetic women the only abnormality found in fetuses and after birth was mild asymmetric septal hypertrophy in 4 cases out of 12 (33%). Three out of 5 in fair control diabetic women showed increased right and left ventricle wall thicknesses (RVW and LVW) and interventricular septum thicknesses (IVS) as well as elevated Right ventricle pre-ejection period/right ventricle ejection time ratio (RPEP/RVET ratio) was consistent with the development of respiratory distress after birth in 3 out of 5. The 3 cases of poor control diabetic women had elevated REW, LVW and IVS thicknesses. Higher than normal in their fetuses and after birth. Two out of the 3 showed elevated Left ventricle pre-ejection period/left ventricle Ejection time ratio (LPEP/LVET ratio) higher than normal in their fetuses and after birth which was consistent with the development of congestive heart failure after birth in them. Elevated RPEP/RVET ratio higher than normal was present in one fetus and after birth in the poor control diabetic woman and this was consistent with the development of respiratory distress after birth in him. There was significant relationship between maternal Glycosylated hemoglobin levels during pregnancy and the cardiac function as well as the clinical severity of cardiopulmonary problems. In conclusion, the degree of glycemic control in pregnant diabetic women affects cardiac function in their fetuses and after birth as well as the clinical severity of cardiac problems postantally.
Source: Molecular Diagnosis and Vaccines 1, pp 95 –101 (2003)More Less
Pityriasis rosea (PR) was long thought to be due to an infectious agent, however, this agent is not known. Recently, the association between PR and human herpesvirus type 7 (HHV7) has been a matter of debate. The work was initiated to investigate if there is an association between HHV-7 and PR with its clinical variants. Thirty patients with PR (17 classical type, 7 without herald patch, and 6 inverse PR), 15 matched dermatological controls (8 tinea corporis and 7 psoriasis), and 15 apparently healthy controls were examined for HHV-7 DNA in plasma by nested polymerase chain reaction (nPCR). Seventeen (56.7%) of PR patients were positive for HHV-7 in plasma, while none of the controls (dermatological and healthy) were positive, (P<0.001). Out of the 17 classic PR patients15 (88.2%), and 2 out of the 7 with secondary rash only (28.6%) were HHV-7 DNA positive, while all inverse PR patients were negative (P< 0.001) in conclusion, PR, especially the classic type, is associated with active replication of HHV-7, which could by primary infection or reactivation of latent infection.
Characterization of methicillin - resistant Staphylococcus aureus isolates from patients at a major hospital in Doha, QatarSource: Molecular Diagnosis and Vaccines 1, pp 103 –110 (2003)More Less
Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important causes of hospital infections worldwide. The mecA gene causes high-level resistance to methicillin. Detection of the MRSAspecific gene using mecA polymerase chain reaction (PCR) has become an important tool for rapid detection of MRSA. Molecular typing of MRSA is crucial for controling the spread and knowing the source of infection. In this study, twenty S. aureus isolates were subjected to mecA PCR then genotyped by pulsed field gel electrophoresis (PFGE) using BamHI restriction enzyme. A 100% correlation was found between the PCR and standard agar plate methods for the detection of MRSA and the 533bp product of mecA amplification was detected in all isolates despite the difference in the type of specimens. PFGE yielded 14 different patterns. 12 isolates made six clusters, each cluster consisted of one pair of isolates while the remaining 8 isolates were non-clustered. Out of the six clusters, three showed more than 80% similarity, while the remaining three pairs showed less than 60% similarity of their restriction fragments. It is concluded that the mecAPCR is a rapid and inexpensive method for diagnosis of MRSA and that PFGE provides a useful tool, being able to determine the relatedness between MRSA. The presence of un-related multi -resistant MRSA strains in the hospital supports the hypothesis that horizontal transfer plays an important role in the dissemination of the mecA gene in the S. aureus population.
Drug resistant Mycobacterium tuberculosis In Qatar: comparison between the conventional and DNA-based methodsSource: Molecular Diagnosis and Vaccines 1, pp 111 –118 (2003)More Less
Early detection of drug resistance in Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium bovis (M. bovis) isolates is crucial for appropriate treatment to prevent the development of further resistance and the spread of resistant strains. The identification of resistance mutations enables the development of molecular tests, which may result in a reduction of turnaround times for susceptibility results to 1 to 2 days. The aim of this study was to estimate the rate of M. bovis within the M. tuberculosis complex (MTC) isolates from Qatar. A second goal was to identify drug resistance to isoniazid (INH), ethambutol (EMb), rifampin (RIF), and streptomycin (STR) by the BACTEC method and mutations in rpoB and katG genes associated with RIF and INH respectively by the Single Strand Conformational Polymorphism (SSCP) method. The study was conducted on 100 MTC isolates from pulmonary tuberculosis patients at Al-Rumela Hospital, Doha Qatar. An allele-specific PCR amplification method was used to differentiate M. tuberculosis from M. bovis cultures. All isolates were found to be M. tuberculosis, and none of them was M. bovis. The conventional drug susceptibility results indicated that only one (1%) isolate is multi drug resistant (MDR) (resistant to both INH and RIF). Drug resistance was demonstrated in 6, 3, 1, and 1 isolates for INH, STR, RIF and EMB respectively. On the other hand, mutation was detected in 4/6 and 1/1 of the INH and RIF resistant isolates respectively. Mutation in the rpoB gene was also detected in two RIF susceptible isolates. It is concluded that M. tuberculosis (not M. bovis) is the main causative agent of human tuberculosis in Qatar and that the rate of drug resistance in M. tuberculosis is very low compared to other countries. The SSCP is a rapid, relatively inexpensive and easy to perform technique.