Tinea unguium diagnosis by polymerase chain reaction [single and multiplex] versus conventional methods

Background: onychomycosis is a common nail disorder, affects approximately 5% of the population worldwide and represents 50% of onychopathies and about 30% of mycotic cutaneous infections. Onychomycosis requires administration of antifungal agents for long periods. Several nail disorders may mimic to onychomycosis clinically. Therefore sensitive, quick and inexpensive test is essential for screening nail specimens. In our labs we commonly diagnose onychomycosis using potassium hydroxide [KOH] preparation and culture. These tests are either time-consuming or require specially trained personnel. A recently developed polymerase chain reaction [PCR] assay has the potential to provide a quick and inexpensive method for diagnosis of onychomycosis. We studied all these different methods in the diagnosis of onychomycosis with special concern on tinea unguium diagnosis

Aim: the aims of this study were to compare KOH preparation, calcofluor white stain [CFW], culture and PCR in the diagnosis of onychomycosis and to determine their sensitivity, specificity, positive predictive value and negative predictive value

Methods: nail specimens were collected from patients clinically suspected of onychomycosis. Nail specimens were examined by KOH 20%, CFW microscopic examination, cultured on Sabouraud's dextrose agar [SDA] containing chloramphenicol, SDA containing cyclohexamide and chloramphenicol and dermatophytes test media [DTM] and lastly nail specimen were subjected to DNA extraction and PCR which was carried out using pan-fungal, pan-dermatophyte, Trichophyton rubrum specific primers

Results: of the 76 patients, 54 had at least I of the 4 diagnostic methods positive for the presence of organisms. Culture was positive only in 46 [46.9%] of nail samples. The most common isolated organisms were Candida albicans [3 6. 9%] followed by T. rub rum [I 7.4%]. Using culture as gold test, sensitivity of KOH, CFW and PCR was 85%, 100%, and 100% respectively while specificity of KOH, culture and PCR was 69%, 67%, and 63% respectively. Conclusion: the application of PCR technology directly to the clinical specimens will allow early and accurate diagnosis of onychomycosis

Levels of Interleukin 17, Fas and eosinophil apoptosis in bronchial asthma patients

Bronchial asthma is a chronic inflammatory disorder of the airway in which many cells and cellular elements play a role. The chronic inflammation causes an associated increase in airway hyper responsiveness that leads to recurrent episodes of wheezing breathlessness, chest tightness and coughing particularly at night or in the early morning. These episodes are usually associated with widespread but variable airway obstruction that is often reversible either spontaneously or with treatment.IL-17 family members belong to a distinct category of cytokines that coordinate local tissue inflammation by inducing the release of pro-inflammatory and neutrophil-mobilizing cytokines. The importance of the IL-17 family in inflammatory and autoimmune disease is becoming increasingly apparent. In this prospective study, we measured sputum IL-17 and serum sFAS in bronchial asthma patients of various disease severities by ELISA technique and also detected sputum eosinophils apoptotic ratio [AR].This study was carried out at Mansoura University Hospitals, Medical Microbiology and Immunology and Thoracic Medicine Departments from August 2007 to August 2009. Fifty bronchial asthma patients and twenty healthy non-smoker subjects were enrolled in the study after informed consent. Sputum IL 17 and serum sFas were measured using ELISA technique and eosinophils apoptotic ratio was detected by identification the morphological features of apoptosis after staining by Giemsa stain. The levels of sputum IL 17 and serum sFAS were increased in bronchial asthma patients especially in severe asthma than control group with statistically significant difference, there is decrease in eosinophils apoptotic ratio in bronchial asthma patients than healthy controls. Bronchial asthma patients have higher levels of sputum IL 17, serum sFAS and decreased eosinophil apoptotic ratio, higher levels of IL 17 were associated with severity of the disease, high levels of serum sFAS inhibit the process of apoptosis and are associated with decrease in eosinophils apoptotic ratio

C-C chemokines, RANTES and eotaxin, in atopic dermatitis, atopic asthmatics and allergic rhinitis patients: relation to disease severity

Allergic asthma is characterized by airway hyper responsiveness and inflammation with tissue and bronchial infiltration by activated eosinophils, T cells, mast cells, and macrophages. The extensive infiltration of eosinophils into the lung is not only a hallmark of allergic asthma but also contributes too much of the damage of respiratory epithelium during late phase airway responses. There is accumulating evidence that chemokines, especially the C-C subfamily, are involved in both the migration and the activation of eosinophil and other leukocytes during asthma responses. Chemokines implicated in asthma include regulated on activation, normal T cell expressed and secreted [RANTES] and C-C chemokine eotaxin. Chemoattractants, including eotaxin, and RANTES, lead to characteristic infiltration by eosinophils, basophils, Th2 lymphocytes, and mast cells in chronic allergic rhinitis. Increased contents of the C-C chemokines, RANTES and eotaxin, were demonstrated in the lesional scales of atopic dermatitis [AD]. Twenty nine patients with a physician diagnosis of asthma and positive skin-prick responses to at least one common aeroallergen were allocated to subgroups according to disease severity as severe asthma [n=12] and mild asthma [n=17] groups. Subjects with asthma were further divided into those with respiratory virus infection [n=10] and those where no respiratory virus infection [n=19]. They were compared with control group of 10 nonsmoking subjects with no positive skin-prick response to common aeroallergens or history of respiratory disease. Regarding allergic rhinitis, subjects were divided into severe allergic rhinitis group [n=10] and mild group [n=10]. Another group of allergic rhinitis patients with nasal polyposis were included as nasal tissue group [n=9]. All previous groups were compared with nonsmoking subjects with no positive skin-prick response to common aeroallergens or history of respiratory disease acted as a control group [n=10]. AD patients were classified into 2 groups, severe AD [n=10], mild to moderate AD [n=10], and a third group [n=10] of apparently normal persons as controls. Scales were collected from skin lesions of AD patients, who had no obvious lesions in their sole skin [n=10]. The mean values of Eotaxin, RANTES, TNF-alpha, eosinophils in severe asthma [70.77 +/- 40.17 pg/ml, 31.27 +/-51.64 pg/ml, 55.36 +/-23.94 pg/ml, 8.91 +/-3.39] are statistically higher than in mild asthma [10.38 +/-7.24 pg/ml, 3.82 +/-1.77 pg/ml, 17.98 +/-10.2 pg/ml, 6.41 +/-2.89] and the mean values of all these parameters were higher in both groups compared to control group [1.88 +/-1.11 pg/ml, 1.14 +/-0.47 pg/ml, 1.30 +/-0.44 pg/ml, 0.0 +/-0.0]. Among 29 allergic asthma patients, 10 patients were infected with respiratory viruses [34.48%], 5 patients with severe asthma and 5 patients with mild asthma. Seven of them were infected with Rhinovirus [24.13%]. The mean values of Eotaxin, RANTES, TNF-alpha, eosinophils in allergic asthma with respiratory virus infection were [43.70 +/-50.33 pg/ml, 27.84 +/-58.85 pg/ml, 39.23 +/-39.23 pg/ml, 7.1 +/-3.72] and comparing them to those allergic asthma without respiratory infection [mild and severe] [30.99 +/-33.64 pg/ml, 8.52 +/-7.74 pg/ml, 30.41 +/-21.86 pg/ml, 7.63 +/-3.14], there were non statistically significant difference. The mean values of Eotaxin, RANTES, TNF-alpha in severe allergic rhinitis [33.6 +/-11.07 pg/ml, 72.17 +/-87.61 pg/ml, 25.47 +/-4.04 pg/ml] are statistically higher than in mild allergic rhinitis [9.80 +/-6.79 pg/ml, 10.50 +/-6.90 pg/ml, 12.99 +/-3.27 pg/ml] and the mean values of all these parameters were higher in both groups compared to control group [0.6 +/-0.69 pg/ml, 0.65 +/-0.74 pg/ml, 0.63 +/-0.54 pg/ml]. The mean values of Eotaxin, RANTES, TNF-alpha in nasal tissue [160.30 +/-51.63 pg/ml, 141.00 +/-55.52 pg/ml, 62.22 +/-7.89 pg/ml] were statistically higher compared to severe [except RANTES], mild allergic rhinitis and control groups. RANTES was not significant between severe allergic rhinitis and nasal tissue due to the wide variations in this chemokine. The mean values of Eotaxin, RANTES, TNF-alpha in severe AD [18.1 +/-9.21 pg/ml, 5.36 +/-2.38 pg/ml, 7.46 +/-0.83 pg/ml] are statistically higher than in mild AD [3.39 +/-1.91 pg/ml, 1.16 +/-0.41 pg/ml, 4.27 +/-1.19 pg/ml] and the mean values of all these parameters were higher in both groups compared to control group [0 +/-0 pg/ml, 0 +/-0 pg/ml, 1 +/-0.66 pg/ml]. The mean values of Eotaxin and RANTES, in severe and mild AD are statistically higher compared to AD [non affected sole] [0 +/-0 pg/ml, 0.3+/-0.48 pg/ml], but there was a non statistically significant difference between AD [non affected sole] and control groups. In conclusion, we emphasized the roles of Eotaxin, RANTES and TNF-alpha at the local level in the pathogenesis of allergic asthma, allergic rhinitis and AD and there relation to disease severity which may direct the attention of therapeutic trials to these locally produced cytokines

Human papillomavirus types 16, 18 DNA, chalmydia antigen and tumour necrosis factor alpha in cervical cancer in women from Dakahlia, Egypt

Infection with human papillomavirus [HPV] and Chlamydia trachomatis are associated with cervical intraepithelial neoplasia. The recognition of HPV infection as a factor that is necessary, but not sufficient, for the development of cervical cancer has resulted in the initiation of several longitudinal studies and randomized clinical trials designed to examine the predictive value of HPV DNA testing. Forty two women were enrolled in this study [patients group] together with 20 apparently normal women [control]. For all patients and control groups, cervical swabs were taken, examined for HPV, HPV 16, HPV 18 DNA by 2 sequential PCR reactions, Chlamydia antigen and TNF-alpha by ELISA. Among 42 cases diagnosed pathologically as cervical carcinoma, HPV DNA was detected in 37 cases [88.09%]. HPV16 DNA was more common than HPV18 DNA as it was detected in 28 cases [66.7%] while HPV18 DNA was detected in 4 cases [9.5%]. There is a statistically significant difference between HPV and control cases, also HPV16 and control cases. Regarding Chlamydia antigen, 10 cases were detected out of 42 cases [23.8%] while, only 3 cases were detected in control group [15%] indicating that there is non statistically significant difference between the two groups. Regarding the pathological types of cervical carcinoma, in adenocarcinoma, HPV DNA was detected in 4 cases [80%], while in Squamous cell carcinoma [SCC], it was detected in 33 cases [89.19%]. In adenocarcinoma, HPV 16 DNA was detected in 2 cases [40%], while in SCC, it was detected in 26 cases [70.27%]. In adenocarcinoma, HPV 18 DNA was detected in 1 case [20%], while in SCC, it was detected in 3 cases [8.11%]. Regarding Chlamydia antigen, in adenocarcinoma, the antigen was detected in 1 case [20%] and in SCC, it was detected in 9 cases [24.32%]. Our results emphasized that HPV 16 was more predominant in squamous cell carcinoma, whereas type 18 was relatively high in adenocarcinoma. The level of TNF-alpha [pg/ml] was 33.81 +/-9.38 in cancer cervix cases, which was statistically significant compared to control cases [1.33 +/-0.74 [P<0.001]]. There is none statistically significant difference between adenocarcinoma [34.12 +/-12.31] and SCC [33.76 +/-9.13 [P=0.970]]. There was no significant difference regarding TNF-alpha level between HPV positive cases [34.23 +/-9.39] and HPV negative [30.66 +/-9.68 [P = 0.342]]. There was a significant difference between HPV16 [32.88 +/-8.84] and HPV18 [41.98 +/-4.32]. Also, there was no significant difference between Chlamydiae positive cases [36.62 +/-8.79] and Chlamydiae negative cases [32.93 +/-9.52 [P=0.260]]. We conclude that cervical infection with HPV but not with Chlamydia may be an important risk factor for the development of cervical cancer and TNF-alpha levels increased significantly in cervical carcinoma without special reference to the pathological type of the tumor