Real-time PCR versus phenotypic diagnostic methods in detection of methicillin and vancomycin resistant staphylococci

Staphylococci are the most frequently isolated bacteria from blood. The prevalence of antibiotic resistance has dramatically increased. Real-time PCR offers rapid, accurate, and sensitive method to detect the presence of antimicrobial resistance. This study is mainly aimed to detect uethicillin [oxacillin] and vancomycin resistant staphylococci isolated from blood of patients with significant bacteremia in Assiut University Hospital using both real-time PCR and phenotypic agnostic methods. Sixty Staphylococcal isolates were included. These isolates were collected from positive blood culture bottles [BACTEC 9050 System] of patients with significant bacteremia in Assiut University Hospital. Identification of staphylococcal species was performed by subculture on Mannitol Salt Agar [MSA] and by Microscan system, while antibiotic sensitivity testing was performed by Microscan system, Epsilometer test [E- test], Disc diffusion [DD] method, Oxacillin Resistance Screening Agar Base [ORSAB] and by real-time PCR for mec A and van A genes. Seven S. aureus isolates and fifty three Coagulase-negative staphylococci [CoNS] were detected by both MSA and Microscan system. The most effective antibiotics for staphylococcal isolates were in order. Vancomycin, Linezolid, Synercid, Rifampicin, Chloramphenicol, Gentamycin, Tetracycline, Trimethoprim/Sulphamethoxazole, Clindamycin, Ofloxacin, Ciprofloxacin, Azithromycin and Erythromycin. Concerning methicillin resistance, Real-time PCR which is the gold standard method detected mecA gene in 57 isolates. Accordingly, sensitivity and specificity of E-test were 964% and 100% respectively, DD method showed 87.7% sensitivity and 100% specificity, ORSAB media showed 92.9% sensitivity and 100% specificity while Microscan showed 100.0% sensitivity and 100.0% specificity. Concerning vancomycin resistance, E-test which is the gold standard method detected vancomycin resistance in 6 staphylococcal isolates. Therefore, the DD method showed 66.7% sensitivity and 100.0% specificity while Microscan showed 83.3% sensitivity and 100.0% specificity. Real-time PCR detected van A gene in only one staphylococcal isolate. CoNS organisms are more implicating than S. aureus in bloodstream infections [BSIs]. About 95% of staphylococcal isolates were resistant to methicillin and 10% were resistant to vancomycin. Real-time PCR was more accurate and rapid method for detection of methicillin resistance than phenotypic methods and it could be considered a confirmatory method for detection of vancomycin resistance in staphylococcal isolates suspected to have the van A gene

Serum Carbohydrate antigen-125 levels as a marker in non Hodgkin's lymphoma

Sixty-seven patients with newly diagnosed non-Hodgkin's lymphoma [NHL] were included in this study. Serial measurements of serum carbohydrate antigen 125 [CA-125] levels were studied in conjunction with serum lactate dehydrogenase [LDH] in monitoring response to treatment. LDH showed increased levels in 52 out of 67 patients. Ten apparently healthy subjects were studied as a control group. A total of 36 out of 67 patients had increased serum CA-125 levels. Increased serum CA-125 levels were significantly associated with advanced stage, higher grade, abdominal serosal involvement, bone marrow involvement and bulky disease. P values were significant in abdominal involvement and in serosal involvement. Elevated serum CA-125 levels were associated with high serum levels of LDH. Of the 36 patients presented with high serum CA-125 levels, 30 patients achieved a complete remission [CR] with normalization of CA-125, four cases achieved partial response and two patients had stable disease and had not achieved normalization of CA-125 levels by the end of the treatment. In conclusion, high serum CA-125 levels were significantly associated with mediastinal, abdominal involvement, high tumor bulk and pleural effusion. Sensitivity of serum LDH and CA-125 was 83.6% and 66.6%, respectively, which indicated that the biologic information provided by them are different. Serum LDH provides a measurement of the tumor load and proliferate activity, while serum CA-125 provides the invasive potential of the tumor

HLA antigens, SICAM-1, INF-Y and hepatitis C virus antibodies in lichen planus

Fifty one patients with lichen planus [LP] were studied for human Leucocyte antigens. [HLA] [class I and II] by micro lymphocyto toxicity assay, serum levels of soluble Intercellular Adhesion Molecule- 1 [sICAM] and Interferon [IFN- gamma] by Enzyme linked Immuno-Sorbent Assay [ELISA] and by Enzyme Amplified Sensitivity Immunoassay [EASIA] respectively, and the detection of Hepatitis C- virus [HCV] anti bodies by Axsym. [ABBOTT laboratories] Seventy two healthy unrelated individuals were studied as control group. There were significant increased frequencies of HLA-A19, B5, CW5, CW7, DR1, and DQ3; a significant decreased frequencies of HLA-CW3, DR4, DQ1 were found in patients in comparison to controls. When patients with mucous membrane affection were compared to controls, there were increased frequencies of HLA-Al9, B5, CW3, CW7, CW17, DR1 and DQ3. Patients without mucous membrane affection showed increased frequencies of B5, CW5, CW7, DR1, DR9, DQ3 and decreased frequencies of HLA-DR4 and DQl in comparison to controls. No significant differences were found when patients with mucous membrane affection were compared to those without mucous membrane affection. No significant differences were also noticed when the clinical types actinic, classic and hypertrophic LP were compared to each other. Sero-positive HCV patients represented 31.4% of the studied LP patients with significant increased frequencies of HLA-B5, CW5, CW7, DR1, DR9 and DQ3 in comparison to controls. Sero-negative HCV patients showed increased frequencies of HLA-B5, CW5, CW7, CW17, DRJ, and DQ3 and significant decreased frequencies of HLA- DR4 and DQ2. No significant differences were found in HLA antigens between Sero-positive and Sero-negative HCV patients. HLA-DRI was found in 90% of patients. This represented a striking statistically significant result in the present study. No significant differences were noticed in serum levels of sICAM-1 and IFN-gamma in patients when compared to controls and when the clinical subgroups were compared to each other. There was no correlation between sICAM and IFN-gamma in the studied patients. The findings of the present study strongly suggest that a genetic predisposition or susceptibility play a role to some extent in the development of LP

Serum immunoglobulins profile, eosinophil count in nasal secretions and blood in nasal allergy with or without bronchial asthma: relationship to bronchial hyperresponsiveness

In this study, the serum levels of IgE, IgA, IgM and IgG in two comparable groups [25 each] of patients with allergic rhinitis with or without asthma, in addition to 20 healthy matching control volunteers were compared. They were also studied for eosinophils count in nasal secretions and blood. These data has been further correlated with lung function and methacholine provocation test of the studied population. The study suggested that there is a high level of blood eosinophils in patients with nasal allergy with or without bronchial asthma. Also, patients with allergic rhinitis with or without asthma pose high serum level of immunoglobulins IgE, IgA, IgM and IgG as well as high level of eosinophils either in blood or in nasal secretions. Lastly, patients with allergic rhinitis even without clinical asthma have air flow limitation and lowPD 20 that positively correlate with blood eosinophil count in patients with concomitant rhinitis and asthma. Hence, the present study objectively demonstrates that nasal allergy and bronchial asthma frequently coexist and they appear to share key elements of pathogenesis with strong physiological and immunological inter-relationship

Evaluation of the role of helicobacter pylori in patients with non-ulcer dyspepsia

This study aimed to evaluate the presence of Helicobacter pylori infection in 36 patients with non-ulcer dyspepsia and the effect of two regimens of short term eradication therapy. All patients were subjected to upper gastrointestinal endoscopy and abdominal ultrasonography. The presence of H. pylori was detected in gastric mucosal biopsy by urease test, gram staining and by culture on campylobacter selective medium. H. pylori was detected in thirty-two patients who were divided into two groups: The first group [twenty patients] received triple therapy of bismuth subcitrate, metronidazole and tetracycline and the second group [twelve patients] received dual therapy with tinidazole and tetracycline. Follow up of patients four weeks after completion of therapy showed that H. pylori was eradicated in 60% of patients in the first group and 45.4% of patients in the second group. Significant improvement as regard symptoms and gastritis was observed after triple therapy. Patients of the first group showed significant improvement of the symptoms of epigastric pain and heart burn, while insignificant improvement was found as regard nausea and vomiting. On the other hand, patients of the second group showed insignificant improvement of the symptoms of epigastric pain, nausea and vomiting, while heart burn remained unchanged. The endoscopic findings were significantly improved after triple therapy in patients in whom H. pylori infection was eradicated, while patients of the second group showed no improvement of gastritis even after treatment and H. pylori eradication. The H. Pylori IgG antibodies were significantly decreased in both groups after treatment regardless of the therapy regimen [in the first group from 136.91 Eu/ml to 102.44 and in the second group from 138.31 Eu/ml to 105.38]

Leucocyte analysis in Coulter STKS in Leucocytosis and malignant blood diseases

Two hundred and thirty-eight selected patients were used to study the leukocyte analysis in the coulter STKS in malignant disease and reactive leukocytosis compared with visual method as a reference method. Group I included 141 patients with leukocytosis [more than 12.0 X 10 9/L] were used to study the total leukocytic count and the differential count. Group II included 97 patients with malignant blood disease and were used to study the suspect flag. It was found that the difference between the results of STKS and that of manual method in the total leukocytic count is highly significant due to multiple technical inherent errors which are liable to occur in counting method. The differential count of the STKS correlated well with the visual methods as regards to lymphocytes, monocytes and eosinophils. A significant difference was found between the results of STKS and visual method as regards neutrophils [underestimation of count] and the basophils [over estimation]

study on HLA-class I antigens in vitiligo: Significant association with A-19

31 unrelated patients with familial vitiligo were examined for a possible association of class I human lymphocyte antigen [HLA] with this disease by microlymphocytic assay. The results were compared with 92 unrelated healthy controls. 9 out of the 31 patients showed a significant increased expression of the antigen HLA-A19 [Pc <0.0001] compared with controls. Extensive vitiligo was marked by a significant increase of HLA-A19 [Pc <0.01], while limited vitiligo showed a significant association with HLA-A3 [Pc <0.05]. Moreover, patients with late age at disease onset showed raised expression of HLA-A3 [Pc <0.05]. These findings suggested that immunogenetic mechanism may be responsible for vitiligo and that unique HLA phenotypes may influence the expression of the disease. A more extensive study of HLA markers in vitiligo patients is necessary before more definitive conclusions can be reached